Vasomotor symptoms, the hot flashes and night sweats that disrupt the lives of millions of women during menopause, are among the most frequent and debilitating symptoms of the transition. They can impair work performance, reduce quality of life, disturb sleep, and drive up healthcare costs. For decades, the mainstay of treatment was menopause hormone therapy, which works well for many women under 60 and within ten years of their final period, but is contraindicated in women with a history of hormone-sensitive cancers, prior blood clots, stroke, or liver disease. Until recently, the only FDA-approved non-hormonal option was the antidepressant paroxetine, while most other non-hormonal drugs were used off-label with modest efficacy and tolerability problems that lead up to half of users to stop within three months.

Neurokinin-targeted therapies changed that landscape. These drugs work by modulating neurokinin receptor activity in the hypothalamus, the brain region that regulates body temperature and becomes dysregulated as estrogen declines. Elinzanetant blocks both NK-1 and NK-3 receptors, and has demonstrated rapid and sustained reductions in the frequency and severity of moderate-to-severe hot flashes in the pivotal Phase III OASIS trials, along with meaningful improvements in sleep disturbance and menopause-related quality of life. But the class has faced scrutiny. Fezolinetant, a selective NK-3 antagonist, has been linked to rare but serious liver injury, prompting an FDA boxed warning for hepatotoxicity and mandatory liver function monitoring in multiple countries. An earlier NK-3 antagonist, pavinetant, had its clinical development halted in 2017 after transient transaminase rises in a Phase II trial.

To build a more comprehensive picture of elinzanetant’s safety, researchers pooled data from four randomized, placebo-controlled trials: the Phase IIb SWITCH-1 study conducted from 2018 to 2019 and the Phase III OASIS-1, -2, and -3 trials conducted from 2021 to 2024. All four enrolled postmenopausal women aged 40 to 65 experiencing moderate-to-severe vasomotor symptoms from natural or surgical menopause. The OASIS-4 trial, which studied women with hormone receptor-positive breast cancer receiving endocrine therapy, was excluded because of its substantially different population. The analysis examined two windows: a placebo-controlled period covering the first 12 weeks, in which 765 women took elinzanetant 120 mg and 754 took placebo, and a full 52-week period including 1,113 women exposed to elinzanetant.

The results showed that half of the elinzanetant group, 50.8 percent, reported at least one treatment-emergent adverse event during the first 12 weeks, compared with 43.2 percent on placebo. Most events were mild or moderate. Headache, fatigue, and somnolence were the most common events, occurring in 7.5, 5.4, and 3.4 percent of elinzanetant users respectively, versus 4.2, 1.3, and 0.5 percent on placebo. Notably, these symptoms clustered in the early weeks: their frequency was the same or lower at 52 weeks than at 12 weeks, suggesting most affected women experienced them only at the start of treatment. Serious adverse events were rare in both groups, at 1.2 percent with elinzanetant and 0.9 percent with placebo over 12 weeks, and nearly all were judged unrelated to the drug. A single serious event, a generalized tonic-clonic seizure in a participant with a long history of epilepsy, was assessed as possibly related; rare seizure cases have since appeared in post-marketing surveillance, and caution is advised for women with seizure disorders.

Discontinuation rates told a similar story of early but limited intolerance. Over 12 weeks, 7.8 percent of elinzanetant users stopped treatment because of adverse events, versus 3.6 percent on placebo, with fatigue and headache the leading causes. Crucially, the discontinuation rate did not rise between 12 and 52 weeks, indicating that women who tolerated the drug initially were unlikely to abandon it later. The authors also noted that because the trials were double-blind, background symptoms of menopause itself, or side effects of permitted concomitant medications, could have been coincidentally attributed to placebo, complicating direct comparisons.

Liver safety was the analysis’s most anticipated finding. Elevated alanine or aspartate transaminases above three times the upper limit of normal appeared in just 0.5 percent of elinzanetant users over 12 weeks and 1.2 percent over 52 weeks, rates that were statistically indistinguishable from placebo. There were no elevations of five or eight times the upper limit during the placebo-controlled period, and no cases meeting the strict Hy’s law criteria that flag potential serious drug-induced liver injury. An independent blinded liver safety monitoring board reviewed all cases meeting close-observation thresholds and judged only one elinzanetant case possibly related, in a woman with a plausible alternative cause involving possible gallstone passage; that case resolved without stopping treatment. The authors point out that elinzanetant lacks the structural features associated with hepatotoxicity and shares no similar substructures with NK-3 antagonists, and that approved NK-1 antagonists have not been linked to liver injury. Most countries do not require liver monitoring with elinzanetant, though the FDA mandates baseline testing and a repeat check at three months.

Cancer risk, a second area of concern raised during fezolinetant’s development, also showed no troubling signal. Malignant tumors were reported in five elinzanetant-treated women, including one case of invasive triple-negative breast cancer judged unrelated to treatment, and two placebo-treated women with basal cell skin cancers. The exposure-adjusted incidence rate for malignant tumors in the elinzanetant group, 0.88 per 100 person-years, was broadly consistent with the population-level rate of 0.81 observed in US women aged 50 to 64 in the SEER cancer registry, and the breast cancer rate of 0.38 aligned with a background rate of 0.29. No cases of endometrial hyperplasia or malignancy emerged in either group, an important finding given that unopposed estrogen in hormone therapy can raise endometrial cancer risk. The researchers also observed that NK-1 receptor antagonists have demonstrated anti-tumor activity in preclinical cancer models, and that elinzanetant modulates neurokinin signaling without any estrogenic effect, offering no mechanistic route to hormone-related cancers. Metabolic markers including blood pressure, cholesterol, triglycerides, and glucose remained stable throughout.

The authors are careful to frame the analysis as exploratory rather than confirmatory. It was not powered for formal hypothesis testing, three of the four trials were placebo-controlled for only 12 weeks, and the enrolled population, which excluded women with recent malignancy and abnormal liver parameters, limits generalizability. The incidence-rate calculations also assume a constant hazard over time, which can mask transient risks, and the comparison with population registry data must be interpreted cautiously given the strict eligibility criteria of clinical trials. Longer-term safety monitoring, time-to-event analyses, and real-world post-marketing evidence will be needed to fully characterize the drug’s profile, and a three-year extension study is underway in the breast cancer population from OASIS-4.

Nevertheless, the pooled data provide the most robust safety picture yet for a treatment that many clinicians view as a watershed for menopause care. For women who cannot or choose not to take hormones, and for the growing number of breast cancer survivors whose endocrine therapy triggers severe hot flashes, the findings support elinzanetant as an effective option with a tolerability profile dominated by mild, early, and self-limited central nervous system symptoms. As post-marketing surveillance accumulates in broader and more diverse populations, the drug class that once seemed jeopardized by liver warnings may be finding surer footing.

Subject of Research: Safety and tolerability of elinzanetant, a dual NK-1/NK-3 receptor antagonist for menopausal vasomotor symptoms, in a pooled analysis of four randomized clinical trials

Article Title: Safety and tolerability of elinzanetant: a pooled analysis from four randomized clinical trials

Article References: Safety and tolerability of elinzanetant: a pooled analysis from four randomized clinical trials. (n.d.). Original publication

Image Credits: AI Generated

DOI: Not provided

Keywords: elinzanetant, menopause, vasomotor symptoms, hot flashes, neurokinin receptor antagonists, liver safety, hepatotoxicity, clinical trials, breast cancer, endometrial safety, pharmacology, women's health

The investigation began with chemistry. Using gas chromatography-mass spectrometry, the researchers confirmed that pulegone dominated the essential oil profile, accounting for 42.72 percent of the mixture. Pulegone is the monoterpene terpene long suspected of causing pennyroyal’s most notorious effects. In the human and animal body, liver enzymes convert pulegone into menthofuran, a proximate toxin that depletes glutathione and destroys hepatocytes, a metabolic pathway first mapped in detail in the 1980s. European regulators at the European Medicines Agency have already issued public statements restricting herbal products containing pulegone and menthofuran, but the precise molecular routes by which the compound harms both liver and reproductive organs have remained fragmented across the literature.

To connect those dots, the team turned to network pharmacology, an approach that maps the interactions between a compound’s chemical constituents and the protein targets encoded in the human or animal genome. By cross-referencing predicted targets of the oil’s constituents with disease pathways linked to hepatotoxicity and reproductive toxicity, the researchers identified 116 shared molecular targets. Among these, three hub genes emerged as the most connected and biologically significant nodes: CYP19A1, the gene encoding aromatase, the enzyme that synthesizes estrogens; ESR2, which encodes the estrogen receptor beta; and HSD11B1, which encodes 11-beta-hydroxysteroid dehydrogenase type 1, a gatekeeper of glucocorticoid action within tissues. All three sit squarely within the steroid hormone biosynthesis and estrogen signaling pathways.

Molecular docking then tested whether pulegone could physically bind these proteins. The simulations returned favorable binding affinities of minus 5.9, minus 6.0, and minus 6.6 kilocalories per mole for CYP19A1, ESR2, and HSD11B1 respectively, values consistent with meaningful ligand-receptor interactions. In other words, the computational analysis suggested that the dominant constituent of pennyroyal oil could directly interfere with the molecular machinery that governs estrogen production, estrogen reception, and intracellular steroid metabolism. This was the hypothesis the animal experiments would go on to test.

For the in vivo phase, the researchers followed OECD Guideline 421, the internationally standardized reproduction and developmental toxicity screening protocol. Wistar rats received pennyroyal essential oil at 100, 200, or 400 milligrams per kilogram of body weight throughout premating, mating, gestation, and lactation, covering the entire reproductive cycle. The design allowed the team to track effects from the earliest stages of gamete development through weaning, capturing both parental toxicity and consequences for the next generation.

The outcomes were dose-dependent and consistent across multiple endpoints. Exposed animals showed impaired fertility, reduced implantation rates, smaller litter sizes, lower pup survival, and slower growth in surviving offspring. Alongside these reproductive failures, the rats exhibited elevated markers of liver injury, evidence of oxidative stress, disrupted hormone levels, and histopathological damage in the liver, testes, and ovaries. The convergence of liver and reproductive harm in the same animals mirrored the computational prediction that a single compound, acting on a shared network of steroidogenic and estrogenic targets, could drive pathology in both organ systems simultaneously.

The mechanistic story that emerges is threefold. First, endocrine disruption: by binding aromatase and estrogen receptor beta, pulegone appears to interfere with the synthesis and signaling of the hormones that orchestrate ovulation, spermatogenesis, implantation, and pregnancy maintenance. Second, oxidative tissue injury: the metabolic activation of pulegone in the liver generates reactive intermediates that overwhelm antioxidant defenses, damaging hepatocytes and, by extension, the steroid-producing cells of the gonads, which are themselves highly vulnerable to oxidative stress. Third, downregulation of the steroid hormone biosynthesis signaling pathway, which the authors identify as the central pathway linking the computational targets to the observed in vivo damage.

The novelty of the work, the authors argue, lies in establishing the first multi-scale mechanistic link between specific computational targets and whole-animal toxicity outcomes for pennyroyal oil. Previous studies had documented pieces of the puzzle: earlier research in pregnant rats showed that Mentha pulegium extracts could induce abortion and fetal toxicity through modulation of pregnancy hormones and inflammatory signaling, and other work had characterized the antimicrobial and antioxidant properties of the oil. But no prior study had tied the network-level identification of CYP19A1, ESR2, and HSD11B1 as hub genes directly to fertility, developmental, hepatic, hormonal, and histopathological endpoints measured in the same experimental animals.

The implications extend beyond traditional medicine. Pulegone-containing oils are used as flavoring agents and fragrances, and regulatory agencies worldwide grapple with setting safe exposure limits for phytochemicals whose toxicity profiles are incompletely mapped. The dose-dependent nature of the effects observed here, spanning 100 to 400 milligrams per kilogram in rats, offers quantitative anchors for benchmark dose modeling and interspecies risk extrapolation. The study also demonstrates the value of integrating network pharmacology with guideline-compliant animal testing: the computational screen correctly predicted the pathways that the in vivo work confirmed, suggesting that such hybrid designs could make safety assessments of herbal actives faster and more targeted.

Caveats remain. The findings derive from a rat model at relatively high doses, and translation to human exposure scenarios, particularly the low doses typical of occasional culinary or aromatherapy use, requires careful benchmark dose analysis and toxicokinetic modeling. The oil’s composition also varies with plant chemotype, geography, and extraction method, meaning that pulegone content, and therefore risk, is not uniform across commercial products. Still, for a plant whose abortifacient reputation dates back to antiquity and whose lethal hepatotoxicity has been documented in case reports for decades, the new study provides something the historical record never could: a coherent, target-level mechanism connecting a single dominant molecule to the destruction of both liver tissue and reproductive function. For regulators, clinicians, and anyone tempted by pennyroyal’s folkloric appeal, the message is unambiguous. The oil’s chemistry is not benign, its targets are now known, and its dose-dependent harm to fertility and the liver has been demonstrated across the full reproductive life cycle in a standardized animal model.

Subject of Research: Hepato-reproductive toxicity of pulegone-rich Mentha pulegium essential oil in rats

Article Title: Mechanistic assessment of hepato-reproductive toxicity of Mentha pulegium L. essential oil in rats

Article References: Pattanashetti, L. A., Siddika, S. A., Swathi, D. S., Sumanth, B., Mishra, M., Kulkarni, G. G., Hiremath, S. I., & Patil, S. B. (2026). Mechanistic assessment of hepato-reproductive toxicity of Mentha pulegium L. essential oil in rats. Molecular Biology Reports, 53(1), Article 1691. https://doi.org/10.1007/s11033-026-12842-3

Image Credits: AI Generated

DOI: 10.1007/s11033-026-12842-3

Keywords: pennyroyal oil, Mentha pulegium, pulegone, hepatotoxicity, reproductive toxicity, endocrine disruption, oxidative stress, network pharmacology, molecular docking, OECD Guideline 421, CYP19A1, estrogen signaling

The research, led by Shaon Basu and Markus Ralser at Charité Universitätsmedizin Berlin and the Francis Crick Institute, together with a team at AstraZeneca, focused on degraders of the androgen receptor, a transcription factor that drives treatment-resistant prostate cancer. First-line therapies for prostate cancer typically block androgen receptor signaling with antagonist drugs, but tumors inevitably acquire mutations at the androgen receptor locus, rendering the drugs ineffective and the disease lethal. Degraders promise a way around this resistance by directly destroying the receptor inside tumor cells. Despite this promise, no androgen receptor degrader has yet demonstrated improved efficacy over standard treatment with an acceptable safety profile, echoing the fate of first-generation bromodomain degraders that failed in cancer patients because of dose-limiting toxicity.

The team’s strategy began with a technological foundation: a fast, scalable proteomic workflow capable of monitoring how thousands of proteins respond to drug treatment in a 96-well plate format. HepG2 cells, a well-established model for screening liver toxicity, were cultivated and treated with liquid-handling robots, and their proteins were digested into peptides and analyzed by data-independent acquisition mass spectrometry on two platforms, using scanning SWATH acquisition on a Sciex TripleTOF 6600 and dia-PASEF acquisition on a Bruker timsTOF HT. Neural network-based processing with the DIA-NN software, benchmarked against a hepatocarcinoma-specific spectral library, quantified thousands of proteins per sample with minimal batch effects and low technical variance, making the approach suitable for industrial-scale screening.

To validate the platform, the researchers exposed HepG2 cells to 78 FDA-approved drugs whose targets span human, bacterial and fungal proteins. Forty-seven of these compounds, particularly those with favorable safety profiles, triggered no significant proteomic response. The remaining 31 induced measurable changes, and the proteomes faithfully reflected each drug’s mechanism of action: methotrexate altered less than one percent of the proteome while its primary target, dihydrofolate reductase, ranked among the most upregulated proteins, whereas the chemotherapy doxorubicin reshaped seven percent of the proteome in a pattern consistent with DNA damage. Intriguingly, the dominant proteomic signatures often corresponded to off-target rather than on-target interactions, including disruption of ribosomal translation, endoplasmic reticulum stress and mitochondrial dysfunction. The antifungal clotrimazole, for example, induced broad rearrangements driven by xenobiotic response proteins and mitochondrial impairment despite the absence of its fungal target in human cells. A principal component analysis cleanly separated compounds with narrow therapeutic windows, such as chemotherapeutics and immunosuppressants, from safer drugs.

With the platform validated, the team turned to the central question: what off-target effects do experimental androgen receptor degraders produce? Because HepG2 cells do not express the androgen receptor, any proteomic response to these compounds must reflect off-target engagement. The researchers profiled a library of 204 structurally unique androgen receptor degraders, including clinically advanced candidates such as bavdegalutamide (ARV-110) and an AstraZeneca compound referred to as compound 1, alongside control degraders targeting BRD4, CDK4 and CDK9. The screening campaign generated 2,113 proteomic samples reaching depths of roughly 4,000 proteins each. The results were striking: despite the absence of their intended target, several androgen receptor degraders perturbed up to 57 percent of the proteome, with an average of 19 percent of proteins differentially expressed, a far stronger response than that elicited by the FDA-approved drug library. Notably, degraders targeting proteins actually expressed in HepG2 cells, such as BRD4, produced responses that partly reflected the biology of their targets, confirming that the platform can distinguish on-target from off-target signatures.

The off-target responses clustered according to chemical structure. Degraders recruiting the cereblon E3 ligase, particularly those using phthalimide derivatives and achiral DHU warheads, produced the broadest perturbations, while those recruiting the von Hippel-Lindau ligase produced the weakest. Crucially, the toxicity was not inherent to the recruiter alone: lenalidomide by itself elicited no proteomic response, while other cereblon ligands induced moderate or strong changes. Functional enrichment analysis pointed decisively at the mitochondrial compartment, with pathways involving mitochondrial protein-containing complexes responding in a dose-dependent manner. When the researchers tested whether the compounds impaired respiration by culturing cells in galactose medium, which forces cells to rely on mitochondrial ATP production, several cereblon-recruiting degraders reduced viability in a dose-dependent fashion. Compound 1 showed a half-maximal inhibitory concentration of 0.73 micromolar in galactose medium, while its constituent building blocks, lenalidomide and an androgen receptor-binding analog of enzalutamide, were inactive above 20 micromolar, indicating a gain-of-function toxicity arising from the assembled degrader molecule.

To pinpoint the molecular mechanism, the team applied machine learning to the proteomic data. Univariate linear regression linking protein expression changes to galactose toxicity revealed that nine of the ten best-fitting proteins localized to mitochondria, and four of these were subunits of electron transport chain complex I, the NADH dehydrogenase that initiates the proton gradient driving ATP synthesis. The complex I protein NDUFA5 showed the strongest relationship, with a coefficient of determination of 0.57. A second-pass gradient-boosted decision tree model, refined through Shapley additive explanations analysis and retrained on top features, achieved a cross-validation area under the precision-recall curve of 80 percent, with complex I proteins dominating the feature importance rankings. Biochemical validation followed: compound 1 reduced oxygen consumption in HepG2 cells, and in assays using bovine heart mitochondrial membranes it potently inhibited NADH oxidation through the complex I-III-IV axis with an IC50 of 0.2 micromolar, roughly 100-fold more potent than lenalidomide and 45-fold more potent than enzalutamide, while leaving complex II activity untouched. The degrader, in other words, was a selective complex I inhibitor.

The most consequential step came next. Because the toxicity was not inherent to the recruiter chemistry, the researchers reasoned that proteomics could identify degraders lacking complex I engagement. Their trained models assigned compound 1 a predicted toxicity score of 91 out of 100, comparable to known hepatotoxins such as troglitazone, but attributed far lower scores of 47 and 35 to two close structural analogs, compounds 2 and 3, which differed only in the linker region and carried a 7-methoxy modification on the lenalidomide moiety. Experimentally, the analogs induced no complex I proteomic signature, did not impair mitochondrial respiration, and showed a complex I IC50 of 6.5 micromolar versus 0.2 micromolar for compound 1. In primary human hepatocytes, compound 1 caused acute cell death at 2.7 micromolar, whereas compounds 2 and 3 were tolerated above 20 micromolar. The analogs also lost the unwanted degradation of the cereblon neosubstrates IKZF3 and SALL4, proteins linked to bone marrow and developmental toxicity, likely owing to the methoxy modification.

Safety gains did not come at the expense of efficacy. All three compounds inhibited proliferation of androgen receptor-positive prostate cancer cell lines, but only compound 1 suppressed androgen receptor-negative lines, suggesting improved selectivity for the refined analogs. Compound 3 degraded the androgen receptor as effectively as compound 1 in LNCaP cells and dose-dependently suppressed prostate-specific antigen expression. It also displayed favorable pharmacokinetics, with low clearance in human liver microsomes and rat hepatocytes and 50 percent oral bioavailability in mouse. RNA sequencing confirmed that compound 3 modulated androgen response signaling similarly to enzalutamide while reducing MYC oncogenic signaling and G2/M checkpoint pathways, consistent with cell-cycle arrest, and without perturbing oxidative phosphorylation at any time point.

The decisive test came in vivo. In a C4-2 xenograft model of castration-resistant prostate cancer in immunocompromised mice, three days of oral dosing with compound 3 at 30 milligrams per kilogram reduced androgen receptor protein levels in tumors by more than 70 percent. Over 28 days of treatment, compound 3 achieved 56 percent tumor growth inhibition, compared with just 15 percent for enzalutamide at 50 milligrams per kilogram, and both treatments were well tolerated with no significant body weight loss. The study establishes that systematic, high-throughput proteomics paired with interpretable machine learning can trace unwanted proteomic signatures back to specific chemical features, here a linker region, and guide the rational design of safer, more selective degraders. As polypharmacological modalities such as degraders and antibody-drug conjugates continue to enter the clinic, the authors argue that this proteome-guided framework offers a versatile route to de-risk drug development before candidates ever reach patients.

Subject of Research: Proteome-guided discovery and mitigation of toxicity mechanisms in androgen receptor-targeted protein degraders

Article Title: Proteome-guided drug discovery maps and mitigates therapeutic degrader toxicity

Article References: Basu, S., Yu, J. S. L., Bosak, J., Bagal, S. K., Crafter, C., Fallan, C., Lemke, O., Freiwald, A., White, M. E. H., Vernardis, S. I., Messner, C. B., Michaloglou, C., Pryde, K., Ramos-Montoya, A., Regan, S., Rodrigo-Brenni, M., Roth, S., Ruffilli, C., Mülleder, M., … Ralser, M. (2026). Proteome-guided drug discovery maps and mitigates therapeutic degrader toxicity. Nature Chemical Biology. https://doi.org/10.1038/s41589-026-02347-2

Image Credits: AI Generated

DOI: 10.1038/s41589-026-02347-2

Keywords: proteomics, targeted protein degradation, PROTAC, androgen receptor, prostate cancer, mitochondrial toxicity, complex I, machine learning, drug discovery, hepatotoxicity, cereblon, mass spectrometry

Benzo[a]pyrene, often abbreviated BaP, is everywhere in modern life. It forms whenever organic matter burns incompletely, which means it rises from charcoal grills as fat drips onto hot coals, drifts from cigarette smoke and tailpipes, and accumulates in workplaces such as aluminum smelters, coke ovens and asphalt plants. The International Agency for Research on Cancer classifies it as a Group 1 carcinogen, a category reserved for agents with confirmed cancer-causing activity in humans. Once ingested or inhaled, BaP is metabolically activated by cytochrome P450 enzymes, chiefly CYP1A1 and CYP1B1, into a highly reactive intermediate called BPDE. This compound latches onto DNA to form adducts that can initiate cancer, while the metabolic process itself generates a flood of reactive oxygen species that overwhelm the body’s natural antioxidant enzymes and damage cell membranes, proteins and lipids. Because the liver serves as the body’s principal detoxification organ, it bears the brunt of this chemical assault.

The research team, based at Islamic Azad University in Shahrekord, Iran, set out to test whether dietary intervention could blunt this damage. Seventy male C57BL/6 mice were randomly assigned to fourteen groups of five animals each, with the sample size determined by formal power analysis to satisfy the principles of reduction in animal research. One group received only standard food, another received the olive oil vehicle used to dissolve the chemicals, and a third received 20 milligrams per kilogram of BaP daily by oral gavage for four weeks, a dose previously shown to induce significant oxidative stress and liver and kidney injury in mice without reaching acute lethal levels. A fourth group received vitamin E at 250 milligrams per kilogram, a dose established in earlier studies as antigenotoxic against BaP damage. The remaining groups received chia seed powder mixed into their food at 10, 20 or 30 percent by weight, either alone, alongside BaP simultaneously, or as a two-week pre-treatment before BaP exposure began.

The design allowed the researchers to answer two distinct practical questions. Co-administration tested whether the protective agents could limit damage occurring at the same time as exposure, a scenario relevant to people who cannot avoid contaminated food or polluted workplaces. Pre-treatment tested whether building up the body’s defenses in advance could prevent injury more effectively, a scenario relevant to preventive nutrition. All biochemical analyses were performed in a blinded manner, and the animals were monitored daily for humane endpoints such as excessive weight loss or lethargy, none of which were reached during the study.

The results were unambiguous in one direction and humbling in another. BaP exposure significantly depleted the mice’s total antioxidant capacity and superoxide dismutase activity, raised malondialdehyde, a marker of lipid peroxidation, elevated the liver enzymes AST and ALT, lowered serum albumin, and pushed triglycerides and LDL cholesterol upward. Vitamin E, despite its reputation as a potent fat-soluble radical scavenger, failed to restore antioxidant capacity or superoxide dismutase activity and offered no meaningful protection to the liver enzymes. Chia seeds, by contrast, produced dose-dependent benefits. At 20 and 30 percent dietary inclusion, co-administered chia significantly raised total antioxidant capacity and superoxide dismutase activity while lowering malondialdehyde. Thirty percent chia pre-treatment increased both superoxide dismutase and glutathione peroxidase activities and cut malondialdehyde levels further. Chia co-administration reduced AST at all doses, rescued albumin at the lower doses, lowered triglycerides across the board, and raised protective HDL cholesterol at the higher doses.

The lipid findings were particularly dramatic. BaP exposure drove triglycerides from roughly 1.1 millimolar in controls to 2.44 millimolar, and LDL cholesterol from about 83 to 163 milligrams per deciliter. Chia co-administration at 30 percent brought triglycerides back near baseline while pushing HDL cholesterol to levels well above those of untreated controls. The authors attribute these effects to the seed’s unique biochemical portfolio: alpha-linolenic acid, abundant dietary fiber and a rich array of polyphenols. Mechanistically, chia-derived alpha-linolenic acid appears to activate peroxisome proliferator-activated receptor alpha, boosting fatty acid oxidation, while suppressing sterol regulatory element-binding protein-1, which curbs lipogenesis. Chia components also inhibit fatty acid translocase, known as CD36, limiting fatty acid uptake into tissues, and the seed’s proteins and polyphenols reduce inflammatory signaling through tumor necrosis factor alpha while raising adiponectin.

Why did the whole food outperform the purified vitamin? The researchers argue that BaP’s toxicity is multifaceted, attacking membranes, enzymes, lipid metabolism and DNA simultaneously, and that a single antioxidant molecule simply cannot cover all those fronts. Chia seeds deploy a multitarget strategy: their polyphenols and omega-3 fats act synergistically to neutralize reactive oxygen species directly, and they appear to activate the nuclear factor erythroid 2-related factor 2, or Nrf2, signaling pathway, which switches on the cell’s own antioxidant gene program. Vitamin E, by contrast, may influence CYP1A1 and CYP1B1 activity, but the study suggests this mechanism is insufficient against the scale of oxidative assault that BaP delivers. The authors also note that the vitamin E dose, though previously validated, may have been inadequate for this model, or that the four-week treatment window was too short.

Not every organ responded. BaP raised blood urea nitrogen and creatinine, classic markers of kidney injury, but neither chia nor vitamin E significantly reversed these changes. The researchers suggest that renal protection may require longer treatment durations or different therapeutic approaches, pointing to similar findings with lavender extract against lead-induced nephrotoxicity. The study also has acknowledged limitations: the diets were not isocaloric, body weight and food intake were not formally tracked, tissue-level analyses were not performed, and the potentially synergistic combination of chia and vitamin E was never tested, a question the authors flag as a priority for future work.

The pre-treatment versus co-administration comparison carries a clear public health message. Pre-treatment with 30 percent chia preserved superoxide dismutase and glutathione peroxidase activities and kept malondialdehyde low, suggesting that building antioxidant reserves before exposure is the more effective strategy. For the millions of people exposed to BaP through charred food, secondhand smoke or occupational settings such as coke ovens and aluminum plants, where air monitoring has identified BaP as the dominant carcinogenic risk, the implication is that preventive dietary supplementation matters more than trying to repair damage after the fact. The findings also align with a broader shift in nutrition science away from single-molecule supplements and toward whole-food matrices, whose complex mixtures of bioactive compounds often deliver benefits that isolated nutrients cannot replicate.

Cautious interpretation remains essential. These results come from mice receiving deliberately high doses of both toxin and protective agents, and human trials of chia supplementation in healthy and overweight adults have generally shown only modest changes in lipid profiles and inflammatory markers, likely because those studies lacked a specific toxic challenge. The authors emphasize that clinical and mechanistic studies in humans are needed before dietary recommendations can be made with confidence. Still, the study offers a compelling proof of concept that a common, inexpensive whole food can outperform a flagship antioxidant supplement against one of the world’s most notorious environmental carcinogens, and it suggests that what you eat before exposure may matter as much as the exposure itself.

Subject of Research: Dietary protection against benzo[a]pyrene-induced oxidative toxicity in mice using chia seeds and vitamin E

Article Title: Protective Effects of Chia Seeds and Vitamin E Against Benzo[a]Pyrene‐Induced Toxicity in Male Mice: A Comparison of co‐Administration and Pre‐Treatment Strategies

Article References: Rezaei, M., & Karimi‐Dehkordi, M. (2026). Protective Effects of Chia Seeds and Vitamin E Against Benzo[a]Pyrene‐Induced Toxicity in Male Mice: A Comparison of co‐Administration and Pre‐Treatment Strategies. Food Science & Nutrition, 14(10), Article e72405. https://doi.org/10.1002/fsn3.72405

Image Credits: AI Generated

DOI: 10.1002/fsn3.72405

Keywords: benzo[a]pyrene, chia seeds, vitamin E, oxidative stress, hepatotoxicity, polycyclic aromatic hydrocarbons, antioxidants, mice study, lipid profile, food toxicology, pre-treatment, nutrition

The distinction begins with chemistry. Ashwagandha’s bioactivity is dominated by steroidal lactones called withanolides, but the withanolide profile differs sharply between organs. Comparative HPLC and UHPLC–MS/MS analyses show that leaves accumulate high levels of withaferin A and related epoxide-bearing withanolides, compounds with potent cytotoxic and anti-inflammatory properties. Roots, by contrast, are relatively enriched in withanolide A, withanolide D, and glycowithanolides, the marker compounds used in pharmacopoeial monographs and standardized commercial root extracts. In chemotype studies such as the variety Poshita, withaferin A peaks in field-grown leaves while withanolide A predominates in roots across multiple chemotypes, establishing a consistent gradient between the more cytotoxic leaf chemistry and the so-called tonic-type root chemistry.

That gradient maps onto a long history of differentiated use. Classical Ayurvedic texts describe ashwagandha primarily as a root-based Rasayana, a rejuvenating tonic prescribed as a powder with milk or ghee for stress, debility, insomnia, and neuromuscular disorders. The leaf appears only sparsely in the classical compendia but features prominently in regional and tribal ethnomedicine across India, where it is applied as pastes, poultices, and decoctions for painful swellings, wounds, boils, scorpion stings, and dental pain, and occasionally ingested for fever and metabolic complaints. In other words, tradition itself drew a line between the root as a systemic tonic and the leaf as a more localized, indication-specific remedy, a historical differentiation of expected benefit–risk profiles that modern science is now rediscovering.

Preclinical pharmacology reflects both shared and distinct activities. Root extracts show robust anti-stress, anxiolytic, anti-inflammatory, cardioprotective, and neuroprotective effects in rodent models, mechanisms linked to modulation of the hypothalamic–pituitary–adrenal axis, GABAergic and serotonergic signaling, and the NF-κB and Nrf2 pathways. Leaf extracts, richer in withaferin A and other electrophilic withanolides, display striking anticancer, immunomodulatory, and neuroprotective activity in vitro and in selected animal models, including selective killing of tumor cells and inhibition of NF-κB and STAT3 signaling. In some comparative experiments leaf preparations match or exceed root extracts on antioxidant and antitumor endpoints, but these studies typically employ higher withaferin A exposures over short durations, leaving the translational safety margins for chronic oral use in humans undefined.

The human evidence base is even more lopsided. Nearly all controlled clinical data derive from root extracts, typically at doses of 240 to 600 milligrams per day, and randomized double-blind placebo-controlled trials in adults with stress, anxiety, or poor sleep consistently report improvements in perceived stress, anxiety scales, sleep quality, and cortisol, with favorable tolerability over 6 to 12 weeks. A 12-month open-label study of a standardized root extract at 600 milligrams per day found no serious adverse drug reactions and no signal for cumulative hepatic, renal, thyroid, or hematologic toxicity, and a systematic review covering 30 clinical trials of root preparations reported no serious adverse events across indications ranging from chronic stress to rheumatoid arthritis and male infertility.

Leaf-specific human data are minimal but revealing. A Phase I open-label trial of a pharmaceutical-grade leaf extract known as RH324 in patients with advanced non-small cell lung cancer reported tolerability and preliminary disease stabilization over 28 days, while a separate Phase I study of a leaf-derived preparation in osteosarcoma patients recorded dose-dependent liver enzyme elevations in five of eleven participants. A small number of randomized trials have tested combined root-and-leaf aqueous extracts for chronic stress and resistance training, but these remain limited. The review’s authors stress that this sparse record does not prove the leaf is inherently unsafe; rather, the leaf evidence is insufficient for chronic systemic use, a critical distinction that much of the current regulatory debate has blurred.

Complicating the picture further is adulteration. The US Botanical Adulterants Prevention Program has documented frequent detection of leaf material in commercial products labeled as root or root-only extract, driven by economics: leaves are cheaper, renewable, and richer in withaferin A. Crucially, the program warns that assays based solely on total withanolides cannot distinguish root from leaf, since withanolides occur throughout the plant. Reliable authentication requires orthogonal methods combining HPTLC or HPLC fingerprinting, targeted quantification of withaferin A and quercetin-related metabolites, and DNA-based tools. This matters because some of the toxicological signal in European risk evaluations derives from studies on leaf or whole-plant preparations rather than authenticated root.

The regulatory consequences have been dramatic. In 2020, a literature-based risk assessment by the Technical University of Denmark concluded that a safe intake level for ashwagandha in supplements could not be established, citing potential thyroid and sex hormone effects and possible abortifacient properties, prompting Denmark to ban ashwagandha-containing supplements from 2023. Independent critiques argue the assessment over-weighted limited animal and case-report data, relied on low-quality sources, and conflated root with leaf and whole-plant preparations while overlooking the broader clinical safety dataset on standardized root extracts. Subsequent evaluations by the Dutch RIVM, Germany’s BfR, France’s ANSES, and the UK Food Standards Agency have flagged hepatotoxicity, thyroid, and reproductive concerns, even while acknowledging that much of the adverse-event literature involves products of uncertain composition and plant part.

Pharmacovigilance adds another layer of ambiguity. Published case reports of liver injury in ashwagandha users describe products that were never analytically characterized for plant part, and causality assessments using the RUCAM method reached only possible or probable scores, never definite. One research group has hypothesized that undeclared leaf adulteration, with its substantially higher withanone and withaferin A content, could contribute to hepatotoxicity, providing a plausible mechanistic basis for the case clusters. If some European hepatotoxicity reports in fact reflect exposure to unlabeled leaf or whole-plant material sold as root, then the central regulatory problem is quality and authenticity rather than plant identity alone, and plant-part-agnostic risk assessments risk punishing the wrong ingredient.

The review’s prescriptions are concrete. Monographs and guidance should define ashwagandha for internal use as root or root extract unless leaf or aerial parts are explicitly declared, with separate dossiers and safety data required for leaf products. Regulators should mandate validated orthogonal authentication methods in manufacturing release and market surveillance, treat undeclared aerial-part content as adulteration, and consider a dose-based framework for leaf anchored to withaferin A limitation, analogous to thresholds set for pyrrolizidine alkaloids, once adequate human pharmacokinetic and dose-escalation data exist to derive a safe exposure level. Priority research needs include comparative randomized trials of authenticated root, leaf, and whole-plant extracts with matched chemotyping, comprehensive hepatic, thyroid, and reproductive safety studies for leaf preparations, and a plant-part-stratified meta-analysis of adverse events across all published trials. Until such data arrive, the authors conclude, authenticated root remains the best-characterized material for systemic use, and the case for treating ashwagandha as a plant-part-specific medicine, not a generic herb, is now firmly on the scientific and regulatory agenda.

Subject of Research: Comparative safety, efficacy, and regulation of ashwagandha root versus leaf preparations

Article Title: Plant Part–Specific Differences in Ashwagandha (Withania somnifera): A Comparative Evaluation of Root and Leaf Evidence for Safety and Regulation

Article References: Dutta, A., & Gupta, J. B. (2026). Plant Part–Specific Differences in Ashwagandha ( Withania somnifera ): A Comparative Evaluation of Root and Leaf Evidence for Safety and Regulation. Food Science & Nutrition, 14(10), Article e72364. https://doi.org/10.1002/fsn3.72364

Image Credits: AI Generated

DOI: 10.1002/fsn3.72364

Keywords: ashwagandha, Withania somnifera, withanolides, withaferin A, herbal supplements, hepatotoxicity, adulteration, phytochemistry, regulation, Ayurveda, food safety, clinical trials

The research team, led by Jane Namukobe and Moses Murungi of Makerere University’s Department of Chemistry, set out to fill that gap using Wistar albino rats, the standard mammalian workhorse of preclinical safety testing. Plant material was collected between April and June from Mpanga Central Forest Reserve and Mabira Forest, authenticated at the Makerere University herbarium, and processed into two very different preparations. Aqueous extracts, made by warm maceration in water at 40 degrees Celsius, were reserved for oral toxicity testing because that is how the plants are traditionally consumed. Organic extracts, prepared with a one-to-one mixture of dichloromethane and methanol, were used for dermal testing because topical remedies are typically blended with vehicles such as petroleum jelly. This design choice matters: what is safe to swallow and what is safe to smear are separate questions, and the study treated them as such.

The testing regime followed Organisation for Economic Co-operation and Development guidelines with slight modifications. For acute toxicity, rats received single oral doses of 2000 or 5000 milligrams per kilogram of body weight, or single dermal doses of 8000 or 10,000 milligrams per kilogram. For repeated-dose studies, animals were dosed daily with 200 or 400 milligrams per kilogram orally for 28 days, or 2000 and 5000 milligrams per kilogram dermally for 21 days. Throughout the studies, researchers recorded clinical observations, tracked body weights weekly, and then euthanised the animals for a full workup: blood chemistry covering liver and kidney markers, organ weights, and histopathological examination of the liver, kidneys, heart, stomach and small intestines. Skin irritation was scored with the Draize system, and sensitisation was probed with a modified mouse ear swelling test using tape stripping to heighten immune responses.

The headline finding is deceptively reassuring: no animal died, and no acute toxicity was observed for any extract by either route. Plectranthus caespitosus emerged as the standout performer. Its aqueous leaf extract produced no clinical signs, no changes in body or organ weights, no biochemical abnormalities and clean histopathology even after 28 days of repeated oral dosing. Dermal application caused neither irritation nor sensitisation. This is the first toxicity study ever conducted on this species, and the result aligns with related Plectranthus species such as P. neochilus and P. amboinicus, which have similarly shown wide safety margins in rodents. For a plant already used against skin allergies, the data make a strong case for pushing its active compounds toward antibacterial and skincare applications.

Erlangea tomentosa tells a more troubling story. A single oral dose of its leaf extract, even at the maximum 5000 milligrams per kilogram, caused only transient effects: elevated respiration, reduced mobility and increased urination that resolved within 24 hours. But repeated oral exposure at 400 milligrams per kilogram produced moderate to severe multifocal necrotizing hepatitis, literally patches of dying liver tissue, along with hyperplasia of goblet cells in the small intestine and sloughing of intestinal villi. Remarkably, the liver damage was invisible in the blood chemistry: bilirubin, creatinine and alkaline phosphatase levels remained statistically indistinguishable from controls, and the ALT-to-ALP ratio stayed below 10. The researchers attribute this mismatch to the liver’s known capacity for rapid biochemical recovery once a necrotizing agent is withdrawn, a phenomenon documented in drug-induced liver injury literature. Histopathology, in other words, caught what blood tests missed.

Repeated dermal exposure to E. tomentosa’s organic extract added another layer of concern. Although the extract was classified as a non-irritant with a primary irritation index below 1.5 and provoked no sensitisation in the ear swelling test, animals dosed repeatedly at 200 and 400 milligrams per kilogram developed necrotizing liver hepatitis. Blood chemistry showed significantly reduced ALT, ALP and urea alongside elevated bilirubin, a pattern the authors note can indicate cholestasis or impaired bile clearance, though reduced transaminases can also reflect hepatoprotective effects. The team points to historical reports of the plant’s toxicity to rabbits, in which fatal doses caused abdominal ascites and congestion of the liver, kidneys and lungs. Phytochemical screening offers a plausible mechanism: the extract contains tannins and anthraquinones, both of which have been shown to induce hepatic necrosis at sufficient concentrations, while its saponins may partially counteract that damage.

Psorospermum febrifugum, the stem bark remedy used across tropical Africa for skin diseases and wounds, occupied a middle ground. Acute oral exposure at 5000 milligrams per kilogram increased liver weight and temporarily raised respiration, urination and mobility, all resolving within a day. Repeated oral dosing at 400 milligrams per kilogram reduced both liver and kidney weights and produced a roughly three-fold increase in the AST-to-ALT ratio, a signature that typically signals hepatocellular injury or stress, together with elevated total and direct bilirubin. Yet histopathology of the liver showed no significant lesions, a discrepancy the researchers interpret as either adaptive organ remodelling, the liver’s regenerative capacity masking early damage, or the slow onset of chronic toxicity. The plant’s tannins and saponins, with their opposing effects on liver tissue, may explain why the tissue looked healthy even as enzyme levels flagged distress.

The dermal picture for P. febrifugum was arguably the most clinically significant. Topical application caused no skin irritation or sensitisation, but repeated exposure at 2000, 5000 and 8000 milligrams per kilogram significantly increased bilirubin, creatinine and AST while decreasing urea and ALP, a combination the authors link to drug-induced cholestasis and impaired kidney function. Acute exposure at 8000 milligrams per kilogram also disturbed sodium and potassium levels. The organic extract is rich in steroids, compounds known to induce or aggravate acute kidney injury, chronic kidney disease and glomerular toxicity, providing a chemical suspect for the renal findings. These results are notable because they emerged from skin application alone, demonstrating that a plant remedy can damage internal organs without ever touching the digestive tract, a fact with real implications for how topical herbal medicines are regulated.

The broader lesson of the study is that safety cannot be inferred from tradition, efficacy, or even a single toxicity endpoint. All three plants passed the most visible tests, no mortality, no acute toxicity, no skin irritation, yet repeated exposure revealed liver and kidney liabilities in two of them, and in E. tomentosa the worst damage appeared only under the microscope. The authors recommend that P. caespitosus advance toward development of antibacterial products and skincare formulations, while E. tomentosa and P. febrifugum require further work to isolate and characterise their toxic constituents, with tannins flagged as the most likely culprits. As antimicrobial resistance drives patients and researchers alike back toward plant-based remedies, this study stands as a cautionary benchmark: the dose, the route and the duration of exposure all matter, and a herbal medicine is only as safe as its most thorough toxicological evaluation.

Subject of Research: Oral and dermal toxicity assessment of three Ugandan ethno-medicinal plants used to treat bacterial skin infections, tested in Wistar albino rats

Article Title: Assessment of oral and dermal toxicity of selected ethno-medicinal plants used to treat bacterial skin infections in Uganda using Wistar albino rats

Article References: Assessment of oral and dermal toxicity of selected ethno-medicinal plants used to treat bacterial skin infections in Uganda using Wistar albino rats. (n.d.). https://doi.org/10.1007/s44339-024-00015-5

Image Credits: AI Generated

DOI: 10.1007/s44339-024-00015-5

Keywords: ethnomedicine, toxicology, Uganda, medicinal plants, hepatotoxicity, nephrotoxicity, Wistar rats, skin infections, phytochemistry, OECD guidelines, dermal toxicity, antimicrobial resistance

Fluoride occupies an unusual position in public health. Deliberately added to drinking water in controlled amounts to protect teeth, it also enters the environment through industrial processes ranging from phosphate fertilizer production to aluminum smelting, coal combustion, and the manufacture of glass, ceramics, and bricks. In regions with fluoride-rich minerals, well water can contain concentrations approaching 10 milligrams per liter, far above levels considered desirable. Food contributes as well: vegetables grown in contaminated fields, fish, and especially tea can carry appreciable fluoride loads, with dry tea leaves averaging around 100 milligrams per kilogram. Because the element is non-biodegradable, exposure is essentially continuous for many populations, making the search for practical protective strategies a matter of genuine urgency.

The biological problem with excess fluoride is that it behaves as a pro-oxidant. When fluoride intake overwhelms the body’s antioxidant defenses, free radicals accumulate faster than they can be neutralized, a state known as oxidative stress. This stress attacks the macromolecules of the cell, peroxidizing membrane lipids, damaging proteins, and breaking DNA strands. Previous studies in mice given sodium fluoride in drinking water showed depleted glutathione in the liver, along with DNA strand breaks, micronucleus formation, and the appearance of oxidized DNA adducts such as 8-hydroxy-2-deoxyguanosine. Such damage can push cells toward genotoxicity, somatic mutation, and apoptosis, the controlled self-destruction that follows irreparable injury. The liver, as the body’s principal detoxification organ, and the kidney, which receives roughly a fifth of cardiac output and concentrates circulating toxins, are particularly vulnerable targets.

To test whether Phyllanthus amarus could intervene, the team harvested leaves from Umuosele community in Amai, Nigeria, dried and powdered them, and extracted 500 grams of the material in methanol over 48 hours. The crude extract was then separated by liquid-liquid partitioning into fractions of differing polarity, using n-hexane, diethyl ether, n-butanol, methanol, and water. Yields varied considerably, with the methanol crude extract giving 37.62 percent, the methanol fraction 28.40 percent, and the aqueous fraction only 7.30 percent. The researchers selected the crude extract, the highest-yielding methanol fraction, and the lowest-yielding aqueous fraction for animal testing, dissolving each in 5 percent Tween 80 to a working concentration of 0.11 grams per milliliter.

Safety came first. In a preliminary lethal dose assessment involving sixty rats, no deaths occurred at oral doses from 100 to 1600 milligrams per kilogram of body weight across the fractions and crude extract. Rats receiving the higher doses showed reduced locomotion, but the 400 milligram per kilogram dose was well tolerated and even associated with normal hair growth, so the team adopted it for the main experiment. Thirty male Wistar rats, aged seven to nine weeks, were then divided into six groups of five. One group served as an untreated control, two groups received sodium fluoride alone or with the Tween 80 vehicle, and three groups received sodium fluoride alongside either the aqueous fraction, the methanol fraction, or the crude extract. Sodium fluoride was dosed orally at 25 milligrams per kilogram per day, and all treatments ran concurrently for 28 days before the animals were sacrificed and their tissues analyzed.

The results were striking. Rats exposed to sodium fluoride alone showed significant decreases in the activities of the antioxidant enzymes superoxide dismutase, catalase, glutathione peroxidase, and glutathione-S-transferase, along with depleted levels of reduced glutathione, in both liver and kidney tissue. At the same time, malondialdehyde, the standard chemical marker of lipid peroxidation and membrane damage, rose significantly. Histological examination confirmed the biochemical picture: livers of fluoride-exposed rats showed necrosis, while kidneys displayed detachment of the glomerulus from Bowman’s capsule, clear signs of structural injury consistent with fluoride-induced oxidative stress.

Co-treatment with Phyllanthus amarus reversed much of this damage. In the groups receiving the aqueous fraction, the methanol fraction, or the crude extract, glutathione levels and the activities of all four antioxidant enzymes rose significantly compared with the fluoride-only animals, while malondialdehyde concentrations fell. The authors attribute this rescue effect to the plant’s rich polyphenol content, which can scavenge free radicals and thereby preserve the metal-dependent antioxidant enzymes that fluoride disrupts. Superoxide dismutase converts the superoxide anion into hydrogen peroxide, which catalase and glutathione peroxidase then eliminate, while glutathione-S-transferase conjugates electrophilic toxins for excretion. By keeping this enzymatic chain intact, the extracts appear to have restored the oxidative balance that fluoride had tipped.

Perhaps the most consequential finding concerned DNA. Using a diphenylamine-based fragmentation assay, the researchers measured how much DNA in liver and kidney tissue had been broken into pieces, a sensitive indicator of genotoxic injury. Fluoride-exposed rats showed markedly elevated DNA fragmentation in both organs, but the three treatment groups showed significant reductions. Notably, the crude extract outperformed the methanol fraction, a difference the authors suggest reflects the higher concentration of bioactive compounds present in the unfractionated preparation. The team proposes that the plant’s antioxidants reduce the electrophilic chemicals that would otherwise attack DNA, preventing the multiple lesions that fluoride toxicity can produce.

The study is not without limitations, which the authors acknowledge candidly. Financial constraints prevented characterization of the specific active constituents in the crude extract and fractions, so the precise molecules responsible for the protective effect remain unidentified. The work was also conducted in a small number of animals over a single 28-day window, and findings in rats do not automatically translate to humans. Nevertheless, the researchers argue that Phyllanthus amarus leaves merit further investigation as an affordable and accessible alternative for managing renal and hepatic disorders linked to fluoride and other oxidative stressors, particularly in regions where fluoride contamination of water and food is endemic.

For a plant that traditional healers have long prescribed for ailments ranging from hepatitis and malaria to diarrhea and skin disease, the new data provide a mechanistic footing for at least one of its claimed benefits. If subsequent studies can identify the active compounds and confirm protective effects at realistic exposure levels, a weed that grows freely across the tropics could become a low-cost ally against an environmental toxin that no one can entirely avoid. For now, the message from the Nigerian laboratory is clear: in rats at least, the chemistry of Phyllanthus amarus stands between fluoride and the fragile DNA of the liver and kidney.

Subject of Research: Protective effects of Phyllanthus amarus extracts against sodium fluoride-induced oxidative stress and DNA damage in rat liver and kidney

Article Title: Impact of Phyllanthus amarus on fragmented DNA and antioxidant activity of rats exposed to sodium fluoride

Article References: Ibobo, G. O., Okpoghono, J., & Onyesom, I. (2025). Impact of Phyllanthus amarus on fragmented DNA and antioxidant activity of rats exposed to sodium fluoride. Discover Toxicology, 2(1), Article 2. https://doi.org/10.1007/s44339-025-00017-x

Image Credits: AI Generated

DOI: 10.1007/s44339-025-00017-x

Keywords: Phyllanthus amarus, sodium fluoride, oxidative stress, DNA fragmentation, antioxidant enzymes, lipid peroxidation, malondialdehyde, glutathione, hepatotoxicity, nephrotoxicity, polyphenols, toxicology

The fruit itself is something of a botanical curiosity. It comes in two distinct forms: a bitter wild type loaded with cucurbitacins, triterpenoid compounds that are highly toxic in large quantities, and a non bitter cultivated form that has gained international acceptance and is now grown commercially in countries such as New Zealand and Australia. The non bitter variety is rich in saponins, oily glycosides that foam when shaken with water and carry a mixture of medicinal properties and potential toxicity. Traditional practitioners have used decoctions of the root to relieve postpartum pain and to treat gonorrhea, while the fruit has been investigated for antimicrobial activity against Salmonella Gallinarum. Yet despite this long history of use, the authors noted a striking gap: almost no toxicological data existed for the fruit in mice, the workhorse animal of modern pharmacology.

To fill that gap, the team collected ripe fruits at the National Veterinary Research Institute, had them authenticated by a plant taxonomist at the University of Jos, and processed them into a powder that was extracted with distilled water using cold maceration over 72 hours. The resulting crude aqueous extract was then tested in Swiss albino mice of both sexes under protocols approved by the institute’s Animal Use and Care Committee. The researchers ran two complementary investigations: an acute toxicity test following Lorke’s method, in which mice received single oral doses ranging from 10 to 5000 milligrams per kilogram of body weight, and a 28 day sub acute study modeled on OECD guideline 407, in which one hundred mice were divided into a control group receiving distilled water and three treatment groups receiving daily doses of 200, 400, or 600 milligrams per kilogram.

The acute results were reassuring. Not a single mouse died or showed abnormal behavior at any dose, from the lowest to the highest, meaning the median lethal dose exceeds 5000 milligrams per kilogram. By the classification standards of Clarke and Clarke, natural substances with lethal doses in the range of 5000 to 15000 milligrams per kilogram are considered only slightly toxic, and OECD guidelines place compounds with values at or above 5000 milligrams per kilogram in class 5, the lowest toxicity category. The finding also aligns with earlier work in rats and cockerels, which similarly reported lethal doses above 5000 milligrams per kilogram for both aqueous and methanol extracts of the fruit. In other words, a one time dose of the ripe fruit extract is, by every measure applied here, remarkably benign.

The 28 day study told a subtler story. Body weights remained unchanged across all groups, suggesting the extract did not suppress appetite, an effect the authors attribute to phytochemicals such as alkaloids, flavonoids, and phenols that are known to regulate feeding. But the blood work revealed significant shifts. Red blood cell counts and packed cell volume rose markedly, with the highest dose group reaching a red cell count of 11.10 million cells per liter on day 21, up from 8.15 at baseline, and packed cell volume climbing from 48.54 percent to 65.11 percent. The researchers suggest these changes may reflect the fruit’s abundant iron, riboflavin, niacin, and amino acids, nutrients known to support red cell synthesis, and they speculate the fruit could hold promise as an anti anemia agent. Total white blood cells, by contrast, fell significantly at the highest dose, dropping from 8.84 to 2.66 billion cells per liter by day 21, alongside reduced lymphocyte counts, a pattern the authors interpret as a possible immunosuppressive effect.

The serum biochemistry raised the loudest alarm bells. Levels of aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase, the classic trio of liver enzymes monitored in clinical practice, rose significantly across the treatment groups. Alanine aminotransferase is primarily liver specific and a reliable marker of liver disease, while aspartate aminotransferase, though found in many tissues, is sensitive to hepatic injury. Albumin and total protein declined, globulin climbed, and direct bilirubin fell at the highest dose. Taken together, these changes point toward liver damage or dysfunction, and the authors flag the presence of coumarins in the extract as a plausible culprit, since these compounds have been linked to hepatotoxicity despite their anticoagulant benefits.

Histopathology confirmed what the blood tests implied. Under the microscope, livers from treated mice showed congestion of hepatocytes at every dose tested, and at 400 milligrams per kilogram the tissue displayed slight necrosis, the death of liver cells. The kidneys and intestines, however, remained structurally normal at all doses. That renal resilience was echoed in the chemistry: urea and creatinine, the standard indicators of kidney function and protein metabolism, actually decreased significantly during treatment. The authors suggest that flavonoids and quinones in the fruit, which scavenge reactive oxygen species and promote diuresis, may exert a nephroprotective effect, consistent with earlier reports of kidney protection from methanol extracts of the fruit in cockerels.

The electrolyte data added one more layer of complexity. Serum sodium rose significantly at the highest dose, from 129.76 to 141.36 milliequivalents per liter by day 28, and chloride increased across all treatment groups. Elevated sodium and chloride can signal dehydration or kidney damage, but since the treated mice showed no dehydration and their kidney tissue appeared normal, the researchers point to another explanation: the fruit itself is rich in sodium, with previous analyses reporting roughly 8.9 to 16.7 milligrams of sodium per 100 grams of seed, so dietary intake from the extract may simply have raised circulating levels directly.

The overall verdict is a cautionary tale about the difference between a single dose and sustained use. The aqueous extract of Cucumis metuliferus appears safe within 24 hours, may support red blood cell production, and shows no signs of harming the kidneys, but prolonged administration risks liver injury, a conclusion consistent with earlier reports of hepatic and renal toxicity in cockerels and rats given extended courses of the extract. The authors emphasize that their findings apply to unrefined, crude extracts, whose chemical composition can vary widely, and they call for further research to identify the specific phytochemicals responsible for both the beneficial hematological effects and the hepatic damage. For the millions of people who turn to traditional plant remedies first, the spiky horned melon now comes with a clearer, and more nuanced, safety label: fine in a pinch, potentially harmful as a daily habit.

Subject of Research: Acute and sub-acute oral toxicity of the aqueous extract of ripe Cucumis metuliferus fruit in Swiss albino mice

Article Title: Toxicity assessment of the ripe fruit of Cucumis metuliferus in Swiss albino mice

Article References: Usman, J. G., Gadzama, U. N., Thliza, C. B., Elisha, I. L., Barde, I. J., Onakpa, M. M., Sanni, S., & Ajagbonna, O. P. (2025). Toxicity assessment of the ripe fruit of Cucumis metuliferus in Swiss albino mice. Discover Toxicology, 2(1), Article 5. https://doi.org/10.1007/s44339-025-00021-1

Image Credits: AI Generated

DOI: 10.1007/s44339-025-00021-1

Keywords: Cucumis metuliferus, horned melon, toxicity, hepatotoxicity, Swiss albino mice, acute toxicity, sub-acute toxicity, hematology, liver enzymes, nephroprotection, traditional medicine, aqueous extract

Senescent cells are cells that have permanently stopped dividing but refuse to die. They secrete a toxic cocktail of inflammatory molecules known as the senescence-associated secretory phenotype, or SASP, and even small numbers of transplanted senescent cells can accelerate aging in mice. Conversely, drugs that clear these cells, called senolytics, have extended healthspan and improved age-related phenotypes in animal studies. The landmark dasatinib-plus-quercetin combination, along with fisetin and UBX0101, established proof of concept, but small-molecule senolytics lack precision and have produced disappointing clinical results accompanied by side effects such as diarrhea, anemia, and rash.

Immunotherapy offers a sharper alternative. uPAR, the receptor for urokinase-type plasminogen activator, is broadly expressed on senescent cells and tumor cells but largely absent from healthy young tissue. CAR T cells engineered against uPAR have already reversed senescence-driven pathologies in mice, including liver fibrosis and metabolic dysfunction. Yet the approach carries risks: high doses of uPAR CAR T cells caused hypothermia, weight loss, and elevated cytokines in mice, and CAR T cells, once infused, expand unpredictably from patient to patient, making their toxicity hard to titrate. The team therefore turned to a bispecific T-cell engager, or BiTE, a soluble antibody construct that recruits a patient’s own T cells to target cells and vanishes from the bloodstream within days, allowing dosing to be adjusted or halted at will.

The engineered molecule, named hGFD-CD3, links the GFD domain of human uPA, the natural ligand of uPAR, to an anti-CD3 antibody fragment derived from the clinically tested drug acapatamab, with a non-functional Fc region added to extend its half-life. In laboratory dishes, the engager bound uPAR-positive senescent human lung fibroblasts and CD3-positive T cells simultaneously, killing senescent cells with an IC50 of just 0.015 nanomolar, a potency the authors attribute to the moderate, natural affinity of the ligand-based recognition domain. Young cells and cells treated with a control construct lacking the GFD domain were spared. Treated T cells upregulated the activation markers CD25 and CD69 and released granzyme along with the inflammatory cytokines TNF-alpha and IFN-gamma.

The in vivo story was more complicated, and it is here that the study makes its most consequential discovery. When aged mice received a high dose of the mouse version of the engager, 1 milligram per kilogram, they died within 24 hours; 0.3 milligrams per kilogram caused ruffled fur, anorexia, and a 20- to 30-fold surge in the liver enzymes AST and ALT. Histopathology pointed squarely at the liver. The reason, the team found, is that liver sinusoidal endothelial cells are among the very few non-hematopoietic cells in the body that express uPAR. Unlike the continuous, basement-membrane-backed vessels of the lung and kidney, liver sinusoids are fenestrated and lack a basement membrane, so widespread destruction of their endothelial lining collapses the microcirculation and injures hepatocytes. In uPAR knockout mice, the engager caused no liver damage at all, proving the toxicity was uPAR-dependent. The same hepatotoxicity appeared with uPAR CAR T cells, even in macrophage-deficient NSG mice, overturning the earlier hypothesis that macrophage activation was to blame.

Crucially, that toxicity proved monitorable and avoidable. At 0.1 milligrams per kilogram, a dose that left liver enzymes and tissues unremarkable, weekly mGFD-CD3 treatment for three weeks significantly reduced uPAR, p16INK4a, and p21 expression across tissues of 22-month-old mice, decreased senescence-associated beta-galactosidase staining in fat, liver, and lung, and suppressed SASP gene expression in liver RNA-sequencing analysis. The engager physically bridged T cells to senescent cells in tissue sections, activated peripheral T cells, and reduced uPAR-positive macrophages. Cytokine rises were modest, roughly twofold, far below cytokine-storm thresholds, and returned to baseline after treatment stopped.

The long-term benefits in mice were striking. Three months after starting treatment, circulating SASP factors had fallen, liver and lung fibrosis had eased, glomerulosclerosis was attenuated, and bone remodeling shifted favorably, with fewer osteoclasts and more active osteoblasts. After six months, treated mice showed better grip strength, treadmill endurance, walking speed, and hanging endurance, along with denser skeletal muscle fibers, less trabecular bone loss, a higher lean-mass fraction, healthier subcutaneous fat distribution, lower fasting glucose, and improved glucose tolerance and insulin sensitivity. Serum transaminases remained within normal ranges throughout, and no tissue showed pathological abnormalities.

The team then crossed the species barrier. In 24-year-old rhesus macaques, roughly equivalent to a 70-year-old human, a dose-escalation trial showed no abnormalities at 10 or 20 micrograms per kilogram, while 40 micrograms per kilogram produced anorexia and a 50 to 100 percent rise in AST and ALT, though still within the normal reference range. At the selected 20-microgram-per-kilogram dose given every three days, two of three macaques tolerated treatment without clinical signs; one showed transient anorexia and transaminase elevations that peaked at 108 U/L for AST and 134 U/L for ALT and cleared within three days. Temporary drops in white blood cells, neutrophils, lymphocytes, and blood pressure all rebounded by day three, a pattern consistent with T-cell migration into tissues rather than destructive depletion, and these fluctuations diminished with repeated dosing.

Over 90 days of follow-up, senescent cell burden in subcutaneous fat, measured by beta-galactosidase staining, fell by day 42, and uPAR, p16INK4a, and p21 expression dropped in fat, muscle, and skin before drifting back up by day 90 as senescent cells slowly re-accumulated. Serum SASP levels fell in parallel, and immune-cell gene signatures declined, indicating reduced tissue inflammation. The macaque with the heaviest senescent burden was the one with the strongest transient enzyme and cytokine responses, reinforcing the authors’ proposal that liver function tests can serve as a practical, biomarker-guided safety gauge for uPAR-targeted senolytics. Because senescent cells are rare, non-proliferative, and take months to return, the team argues that intermittent, low-dose regimens, potentially as infrequent as twice yearly, could impose minimal treatment burden. They caution that for preventive anti-aging use the risk-benefit balance demands careful evaluation, and suggest that diseases with clear unmet need, such as chronic kidney disease, Alzheimer’s disease, and liver fibrosis, may be the most sensible first indications for clinical translation.

Subject of Research: Senolytic immunotherapy targeting uPAR with a bispecific T-cell engager in aged mice and non-human primates

Article Title: uPAR‐Targeting T Cell Engager Exerts Senolytic Effects in Mice and Non‐Human Primates With Serum Aminotransferase Activity as a Safety Monitor

Article References: Deng, J., Zeng, X., Guo, J., Li, L., Zou, S., Liu, N., He, S., Yan, L., Wang, Q., Zhu, Y., Wang, B., Yang, D., Lv, L., & Zhao, X. (2026). uPAR ‐Targeting T Cell Engager Exerts Senolytic Effects in Mice and Non‐Human Primates With Serum Aminotransferase Activity as a Safety Monitor. Aging Cell, 25(10), Article e70742. https://doi.org/10.1111/acel.70742

Image Credits: AI Generated

DOI: 10.1111/acel.70742

Keywords: senolytics, cellular senescence, uPAR, T-cell engager, BiTE, aging, hepatotoxicity, aminotransferases, CAR T cells, rhesus macaques, SASP, liver sinusoidal endothelial cells

Mycotoxins are toxic secondary metabolites produced by fungi, and more than 400 different types have been identified from over 100 fungal strains. They contaminate agricultural products through molds such as Aspergillus, Penicillium, and Fusarium, and by some estimates roughly a quarter of the world’s crops, including cereals, pulses, oilseeds, and nuts, are affected by mold and fungal contamination. Humans and animals are exposed mainly through ingestion, though dermal and inhalation routes also matter, and the severity of harm depends on the toxin type, the dose and duration of exposure, nutritional status, and interactions with other chemicals. Because real-world diets rarely contain a single contaminant, the question of how mycotoxins interact inside the body is not academic; it is central to food safety.

Aflatoxin is among the most notorious of these compounds. Produced primarily by Aspergillus flavus, A. parasiticus, A. nomius, and A. aflatoxiformans, the aflatoxin family includes 18 known types, of which AFB1, AFB2, AFG1, AFG2, AFM1, and AFM2 are the most toxic and are classified as Group 1 carcinogens. The liver is the primary target. Aflatoxin B1 is metabolized by microsomal CYP450 enzymes into AFB1-8,9-epoxide, a highly reactive form that binds DNA and proteins and drives tissue toxicity; a characteristic G to T transversion at codon 249 of the P53 gene has been linked to aflatoxin-induced hepatocellular carcinoma. An estimated 28 percent of hepatocellular carcinoma cases worldwide are associated with AFB1 exposure, and acute aflatoxicosis outbreaks have been reported in India, Kenya, and Tanzania.

Ochratoxin A, the second toxin in the study, is produced by filamentous fungi including Aspergillus and Penicillium species and exists in three forms, with OTA being the most toxic and the most frequently detected in food and feed. It exerts a remarkably broad toxicological portfolio: mutagenicity, hepatotoxicity, neurotoxicity, nephrotoxicity, teratogenicity, and immunotoxicity. OTA has been implicated as a possible etiological factor in Balkan Endemic Nephropathy, a mysterious tubulo-interstitial kidney disease seen in the Balkan Peninsula and Romania, and has been associated with chronic interstitial nephropathy in humans as well as renal and urothelial cancers in rats. Unlike aflatoxin, OTA is considered a cumulative toxin, building up in the body through repeated dietary exposure.

To test what happens when these two contaminants meet, the team, led by Kanchan Gopal Choudhary of Patna University together with colleagues at Mahavir Cancer Sansthan and Research Centre, ran a carefully controlled sub-acute toxicity experiment. Adult male Charles Foster rats, roughly eight weeks old and weighing 160 to 180 grams, were randomly assigned to groups receiving aflatoxin at 0.9 milligrams per kilogram of body weight per day, ochratoxin A at 2.75 milligrams per kilogram per day, or a combined half-dose regimen of 0.45 plus 1.375 milligrams per kilogram per day, with untreated controls for comparison. Each treatment arm was subdivided into 30-, 45-, and 60-day exposure periods. The toxins themselves were extracted from toxigenic fungal cultures, aflatoxin from Aspergillus flavus isolated from contaminated stored wheat and ochratoxin A from Aspergillus niger isolated from contaminated sesame seeds, and their concentrations were verified by thin-layer chromatography and high-performance liquid chromatography with fluorescence detection.

The hematological findings pointed to anemia across the single-toxin groups. Aflatoxin-treated rats showed significant drops in red blood cell counts, hemoglobin, platelets, lymphocyte percentage, basophil counts, and packed cell volume, alongside rises in neutrophil percentage, mean corpuscular volume, and mean corpuscular hemoglobin, a pattern consistent with normocytic normochromic anemia. The ochratoxin A group displayed a similar but distinct signature, with reduced red cells, platelets, packed cell volume, lymphocytes, monocytes, and basophils and elevated MCV, MCH, and MCHC, suggesting normocytic hypochromic anemia. The authors attribute these declines to mechanisms including hematopoietic cellular abnormalities, reduced iron-binding capacity, and inhibition of protein synthesis through decreased serum albumin. Notably, the combined-exposure group produced no clear hematological pattern at all, an early hint that co-exposure behaves in unpredictable ways.

Liver chemistry told a story of accumulating hepatic stress. Serum AST, a transaminase released when hepatocytes die, rose significantly in the 60-day groups of all three treatment arms, while alkaline phosphatase climbed significantly in the aflatoxin and ochratoxin A groups. Other liver markers, including ALT, total bilirubin, and direct bilirubin, remained unchanged, indicating that prolonged rather than short-term exposure is what tips the liver into measurable dysfunction. Under the microscope, the damage tracked the biochemistry: aflatoxin-treated livers progressed from cytoplasmic vacuolization and sinusoidal dilation at 30 days to focal necrosis at 45 days and, by 60 days, degenerated cytoplasm, irregular Kupffer cell nuclei, and mild hepatic steatosis. Ochratoxin A and the combination produced comparable trajectories of ballooning hepatocytes, periportal hemorrhage, mononuclear infiltration, and necrosis, with quantitative scoring of degeneration among 100 hepatic cells per animal confirming steadily worsening injury over time.

The kidney results were where the study took its most unsettling turn. In aflatoxin-treated rats, creatinine, a standard marker of renal filtration failure, rose significantly at 30 and 45 days but returned to near-control values by 60 days, a rebound the authors suggest may reflect induction of cytochrome P-450 enzymes such as CYP3A4 that metabolize aflatoxin with prolonged exposure. Ochratoxin A produced the expected nephrotoxic profile, with uric acid elevated at 45 days and creatinine elevated at both 45 and 60 days, confirming dose- and time-dependent renal injury. Histologically, all treated groups showed glomerular degeneration, damage to the brush border of Bowman’s capsule, cytoplasmic vacuolization of the epithelial lining, and degeneration of proximal and distal convoluted tubules, with hemorrhages inside Bowman’s capsule appearing in the combined group by 60 days.

The combination group, however, broke the expected correlation between blood chemistry and tissue damage. Creatinine rose significantly only at 45 days and had normalized by 60 days, while uric acid, urea, and blood urea nitrogen never significantly changed, yet histopathological analysis of the same animals revealed chronic, progressive kidney toxicity, including tubular destruction, glomerular degeneration, and hemorrhage. In other words, the laboratory values suggested recovery or tolerance at precisely the time point when the tissue showed the most severe structural damage. The authors conclude that aflatoxin plus ochratoxin A may act as a silent toxicant, one that evades the standard biomarker panel used to detect kidney injury, and they caution that biochemical normality should not be mistaken for histological health in co-exposure scenarios.

The findings carry real-world weight because co-contamination of food and feed with multiple mycotoxins is the norm rather than the exception, and regulatory limits are typically set toxin by toxin. If half-doses of two common contaminants can quietly destroy renal tissue while leaving blood tests clean, current monitoring strategies may systematically underestimate the health burden of mycotoxin mixtures, particularly in regions where dietary exposure is chronic and healthcare diagnostics rely heavily on serum chemistry. The authors acknowledge a key limitation: toxin levels in the blood were not measured, so pharmacokinetic interactions between aflatoxin and ochratoxin A remain to be characterized. Follow-up studies tracking co-exposure with direct toxin quantification, and ideally with more sensitive kidney injury biomarkers beyond creatinine, will be needed to confirm how generalizable this silent-toxicity phenomenon is, and whether human populations eating contaminated staples face the same invisible risk.

Subject of Research: Sub-acute toxicity of combined aflatoxin and ochratoxin A exposure in Charles Foster rats

Article Title: Synergistic impact of aflatoxin and ochratoxin A exposure in Charles foster rats: a toxicological study

Article References: Choudhary, K. G., Sharfuddin, C., Kumar, A., & Ghosh, A. K. (2025). Synergistic impact of aflatoxin and ochratoxin A exposure in Charles foster rats: a toxicological study. Discover Toxicology, 2(1), Article 16. https://doi.org/10.1007/s44339-025-00036-8

Image Credits: AI Generated

DOI: 10.1007/s44339-025-00036-8

Keywords: aflatoxin, ochratoxin A, mycotoxins, nephrotoxicity, hepatotoxicity, silent toxicity, Charles Foster rats, food safety, normocytic anemia, histopathology, creatinine, toxicology