Erythema nodosum leprosum, often abbreviated ENL, is one of the most feared complications of leprosy. It is an immune-complex mediated condition that produces fever, tender skin nodules, nerve pain, and systemic inflammation, and it can recur in waves over months or even years. Standard treatment relies on high-dose corticosteroids, typically prednisolone, but prolonged steroid exposure carries a heavy burden of adverse effects, including diabetes, hypertension, osteoporosis, weight gain, and increased susceptibility to infection. Clinicians have long searched for a steroid-sparing agent that could control the inflammation while limiting this collateral damage.

Methotrexate emerged as a promising candidate. The drug, which suppresses inflammation by interfering with folate metabolism and is a mainstay of rheumatoid arthritis and psoriasis treatment, is affordable, orally administered, and already familiar to health systems in many leprosy-endemic countries. Earlier studies suggested possible benefit, but the evidence base was thin, consisting largely of small, uncontrolled observations. The ENLIST group, an international consortium of leprosy researchers, designed the MaPs in ENL trial to settle the question with rigorous methodology.

The trial enrolled adults aged 18 to 60 years with severe ENL. Between January 2023 and June 2024, 231 individuals were screened and 137 were randomised. Sixty-eight participants received weekly oral methotrexate at 15 milligrams, increased to 20 milligrams for those weighing 60 kilograms or more, in addition to prednisolone. Sixty-nine received matching placebo plus prednisolone. Critically, all participants received an identical 20-week prednisolone regimen beginning at 40 milligrams daily, ensuring that any difference in outcomes could be attributed to the methotrexate rather than to variations in steroid dosing. Participants were followed for a total of 60 weeks.

The primary outcome was the time to the first ENL flare requiring additional prednisolone, assessed at 24 and 48 weeks. The results were unambiguous. By 24 weeks, 85 of the 137 participants, or 62.0 percent, had experienced a flare requiring extra steroid therapy. The adjusted hazard ratio comparing methotrexate with placebo was 0.98, with a 95 percent confidence interval of 0.62 to 1.54, a result statistically indistinguishable from no effect. By 48 weeks, 102 participants, or 74.5 percent, had flared, and the adjusted hazard ratio was 0.95 with a confidence interval of 0.62 to 1.43. In plain terms, adding methotrexate neither delayed flares nor reduced how often patients needed rescue steroids.

Secondary outcomes told the same story. Methotrexate did not reduce the severity of ENL at the first flare, the frequency of flares, or the severity of subsequent episodes. Health-related quality of life, measured throughout the trial, improved substantially in both groups, but there was no evidence of a differential treatment effect between the methotrexate and placebo arms. On a more positive note, methotrexate was generally well tolerated, with no signal of serious safety concerns emerging from the trial data. The study was registered at ClinicalTrials.gov as NCT03775460 on 29 November 2018, and the trial’s methodological transparency, including its double-blind design and pre-specified outcomes, strengthens confidence in the null result.

The implications for clinical practice are significant. For decades, the management of severe ENL has been one of the most challenging aspects of leprosy care, and the prospect of a steroid-sparing option generated genuine enthusiasm among specialists. Methotrexate’s low cost made it especially attractive for the low-resource settings where leprosy remains endemic. The trial’s findings now mean that clinicians and guideline developers should not expect methotrexate to reduce steroid dependence in these patients, and the study’s authors state plainly that their results do not support the use of methotrexate for severe ENL.

Yet the trial also delivers valuable knowledge beyond the negative headline. It demonstrates that large, high-quality randomised trials are feasible in leprosy-endemic settings, spanning four countries and multiple referral centres with rigorous blinding and follow-up. It provides some of the best available natural history data on ENL, showing that nearly three-quarters of patients with severe disease experience flares requiring additional steroids within a year even under a standardised treatment protocol. This underscores how aggressive and persistent the condition can be, and it establishes a benchmark against which future therapies can be measured. The finding that quality of life improved substantially in both groups also offers a measure of reassurance that the standard prednisolone regimen, whatever its side-effect burden, delivers meaningful symptomatic relief.

The search for better ENL treatments now turns elsewhere. Researchers have investigated other immunomodulatory agents, including thalidomide, which remains effective in some settings but is restricted or unavailable in many countries because of its teratogenic history, as well as newer biologic approaches that remain largely untested in this population. The MaPs in ENL trial, by rigorously excluding methotrexate, helps direct attention and funding toward candidates with a stronger mechanistic rationale. For the estimated hundreds of thousands of people affected by leprosy worldwide, and for the minority who develop severe ENL reactions, the trial is a sobering reminder that progress against neglected tropical diseases often advances one carefully conducted negative result at a time. The full dataset and the collaboration behind it, coordinated through the Erythema Nodosum Leprosum International STudy Group, now constitute a foundation on which the next generation of ENL trials can be built.

Subject of Research: A randomised controlled trial of methotrexate as a steroid-sparing treatment for severe erythema nodosum leprosum

Article Title: Methotrexate and Prednisolone compared to placebo and prednisolone in the treatment of severe Erythema Nodosum Leprosum: An international multicentre, double-blind randomised controlled clinical trial—MaPs in ENL

Article References: de Barros, B., Sultana, F., Maximus, N., Pai, V. V., Wakade, A., Bhame, B., Acharya, B., Hamza, A., Getachew, A., Alinda, M. D., Listiawan, M. Y., Doni, S. N., Hagge, D. A., Napit, I., Shah, M., Darlong, J., Nicholls, P., Genser, B., Lambert, S. M., … on behalf of the Erythema Nodosum Leprosum International STudy (ENLIST) Group (2026). Methotrexate and Prednisolone compared to placebo and prednisolone in the treatment of severe Erythema Nodosum Leprosum: An international multicentre, double-blind randomised controlled clinical trial—MaPs in ENL. PLOS Neglected Tropical Diseases, 20(9), e0014739. https://doi.org/10.1371/journal.pntd.0014739

Image Credits: AI Generated

DOI: 10.1371/journal.pntd.0014739

Keywords: leprosy, erythema nodosum leprosum, methotrexate, prednisolone, randomised controlled trial, neglected tropical diseases, clinical trial, immunomodulation, corticosteroids, ENLIST, PLOS Neglected Tropical Diseases, steroid-sparing therapy

The research team, led by Jenny Linnea Victoria Lindroos of the University of Bergen and the Norwegian Institute of Public Health, exploited a unique strength of the Nordic healthcare infrastructure: mandatory national registers that record every birth, every hospital contact, and every prescription dispensed at a pharmacy. By harmonizing and pooling data from the Norwegian medical birth register covering 1999 to 2023 and its Swedish counterpart covering 1994 to 2022, and linking these records individually through each mother’s national identity number, the investigators assembled a cohort of 339 pregnancies among women with confirmed myasthenia gravis, drawn from more than 2.5 million births. The diagnosis itself was verified through multiple routes, including hospital diagnoses coded under the International Classification of Diseases and repeated fills of pyridostigmine, the standard symptomatic treatment, with validation studies in Norway showing a positive predictive value of 99 percent for the register-based diagnosis.

The methodological innovation at the heart of the study was group-based multi-trajectory modeling, an unsupervised statistical clustering technique that can follow several medication classes simultaneously across repeated time windows. Rather than imposing predefined categories such as early versus late pregnancy, the model let the data speak, dividing the timeline into four pre-pregnancy quarters, three trimesters, and two postpartum periods, and estimating how each pregnancy’s use of symptomatic drugs, oral corticosteroids, and non-steroidal immunosuppressants evolved together. The algorithm tested three, four, and five latent groups, ultimately settling on a four-group solution that balanced statistical fit, measured by an entropy of 0.92 and posterior probabilities between 95 and 99.6 percent, against clinical interpretability and privacy constraints that ruled out groups smaller than ten pregnancies.

Among the 339 pregnancies, 201, or roughly 59 percent, involved at least one fill of a myasthenia gravis medication during the year before pregnancy or during pregnancy itself, while 138 women, about 41 percent, filled no myasthenia gravis prescriptions at all across that entire window. The four trajectories that emerged were named for their defining patterns: Intermittent users, the largest group at 20.9 percent of the cohort; Persistent symptomatic-only users at 19.8 percent; Immunosuppressant plus symptomatic users at 10.3 percent; and Immunosuppressant-only users at 8.3 percent. The two largest groups relied exclusively on symptomatic medications such as pyridostigmine but differed sharply in consistency. Persistent symptomatic-only users refilled their prescriptions in 75 to 88 percent of every time period, averaging two fills per period, whereas Intermittent users filled prescriptions in only 21 to 28 percent of pre-pregnancy periods and just 13 percent during the second trimester. The average dispensed pyridostigmine dose told the same story, at 269 milligrams per day for the persistent group versus 159 for the intermittent group.

The immunosuppressant groups displayed more complex dynamics. Immunosuppressant plus symptomatic users carried the heaviest treatment burden, purchasing pyridostigmine at an average of 450 milligrams per day during the first trimester and prednisolone at roughly 16 milligrams per day, double the dose seen in the immunosuppressant-only group. Strikingly, their use of non-steroidal immunosuppressant therapy, chiefly azathioprine, fell steeply after conception, from around 60 percent filling prescriptions before pregnancy to only 20 percent in the third trimester. The authors interpret this drop as a deliberate attempt to avoid fetal drug exposure, though they cannot exclude improvement in disease activity, side effects, or pregnancy-related vomiting as contributing causes. In contrast, Immunosuppressant-only users maintained a remarkably stable prednisolone trajectory of about 8 milligrams per day throughout the entire study period, below the 10-milligram threshold generally considered safe in pregnancy, and rarely needed symptomatic medications at all.

Safety signals in the dispensing data were reassuring in several respects. Known teratogenic drugs, specifically methotrexate, mycophenolate mofetil, and cyclophosphamide, were never dispensed later than nine to twelve months before pregnancy, indicating that clinicians and patients successfully cleared these hazardous agents well before conception. Tacrolimus was never refilled during pregnancy, and prednisolone was the only oral corticosteroid used throughout. Biologics such as rituximab appeared only in pre-pregnancy and postpartum windows, and plasma exchange was confined to the pre-pregnancy period. Azathioprine, considered acceptably safe based on observational and animal data, was the dominant non-steroidal immunosuppressant, and its use actually rose during the final nine months before conception among the heaviest treatment group, a pattern the authors read as evidence of preconception treatment optimization.

Hospital contact patterns added a crucial clinical dimension to the medication trajectories. Overall, myasthenia gravis admissions were infrequent, but they clustered conspicuously in one group: 23 percent of Immunosuppressant plus symptomatic users were admitted for myasthenia gravis during pregnancy, and 26 percent had at least one such admission before pregnancy, with 71 percent requiring myasthenia gravis outpatient visits. This same group also showed the highest body mass index, more diabetes and gestational hypertension, and, tellingly, the lowest use of preconception folic acid supplementation, with fewer than five women taking it. Combined with the observation that 23 percent of these women had no specialist outpatient contact at all in the year before pregnancy, the authors argue that many of these pregnancies were likely unplanned and not preceded by neurological counseling, a gap with tangible consequences given that this group also frequently stopped immunosuppression during gestation.

By contrast, the Non-users, who made up 41 percent of the cohort, almost never required myasthenia gravis admissions during pregnancy or postpartum and rarely initiated treatment after delivery, consistent with stable remission or minimal disease manifestation. The Immunosuppressant-only users presented a paradox: minimal myasthenia gravis hospital contacts but frequent non-myasthenia specialist care, likely reflecting the known associations of corticosteroid exposure with hypertension, hyperglycemia, infections, preeclampsia, and preterm birth, alongside a high rate of autoimmune comorbidity at 39 percent and assisted reproductive technology conception in 21 percent of the group. Notably, the trajectory groups aligned closely with the Myasthenia Gravis Foundation of America Post-intervention Status classification, suggesting that dispensing records alone can proxy disease severity classes without requiring clinical examination data.

The study’s limitations deserve honest framing. Infusion therapies such as rituximab, administered in hospitals rather than pharmacies, were invisible to the prescription registers in Sweden and in Norway before 2022, and the drug’s long-lasting immunosuppressive effect may partly explain the large Persistent symptomatic-only group, particularly its Swedish predominance. Spontaneous and induced abortions could not be included because birth registers incompletely capture them, meaning the findings generalize best to pregnancies progressing to birth, and severe cases may be underrepresented. The authors also lacked antibody status, thymectomy history, and formal severity scores. Even so, the cohort’s population basis, prospective data capture, and cross-national harmonization make it an unusually robust evidence base.

The practical message for clinicians is unambiguous. Most women with myasthenia gravis enter pregnancy with no, low, or stable medication use and remain on predictable trajectories, but the minority who combine immunosuppressants with symptomatic therapy face the highest risk of disease exacerbation, hospitalization, and abrupt treatment discontinuation precisely when stability matters most. The authors conclude that identifying these trajectory subgroups, which likely represent distinct disease courses in relation to pregnancy, could enable individualized care pathways, and that preemptive discussions about reproductive plans should be routine when treating women of childbearing age with myasthenia gravis. For a disease whose course in pregnancy has long been described as unpredictable, this register-based cartography of real-world drug use turns a page: the unpredictability, it turns out, has structure, and that structure can now be named, measured, and planned around.

Subject of Research: Medication use trajectories during pregnancy in women with myasthenia gravis

Article Title: Medication use trajectories in myasthenia gravis pregnancies: a population-based drug utilization study

Article References: Lindroos, J. L. V., Gilhus, N. E., Brauner, S., Bjørk, M.-H., Midelfart-Hoff, J., Cesta, C. E., Furu, K., & Cohen, J. M. (2026). Medication use trajectories in myasthenia gravis pregnancies: a population-based drug utilization study. Journal of Neurology, 273(10), Article 572. https://doi.org/10.1007/s00415-026-14115-2

Image Credits: AI Generated

DOI: 10.1007/s00415-026-14115-2

Keywords: myasthenia gravis, pregnancy, pharmacoepidemiology, pyridostigmine, prednisolone, azathioprine, immunosuppression, trajectory modeling, Nordic registers, autoimmune disease, neuromuscular disorders, maternal health

The review, conducted by a team at Manipal Academy of Higher Education and registered prospectively with PROSPERO, followed the PRISMA 2020 reporting guidelines and searched PubMed, Scopus, and Embase for studies published between June 2015 and June 2025. From 975 initial records, the researchers whittled the field down to 105 eligible studies, all of which used chemicals to induce osteoporosis-like bone loss in zebrafish. The sheer volume of work is striking: publications rose steadily over the decade, with a notable surge after 2021 and a peak of 24 studies in 2024 alone, signaling rapidly growing interest in the tiny striped fish as a skeletal research platform.

Why zebrafish? The answer lies in a remarkable combination of biology and practicality. Humans and zebrafish share more than 70 percent of their genes, with at least one zebrafish ortholog existing for the vast majority of human genes. Crucially, the molecular machinery that governs bone remodeling, including the Wnt/beta-catenin, BMP, and RANK/RANKL pathways, is conserved between fish and mammals. Zebrafish larvae are optically transparent, which means researchers can watch bone mineralization unfold in real time under a microscope using fluorescent dyes, without sacrificing the animal. Their small size, rapid development, and high fecundity make them dramatically cheaper and faster than mice or rabbits, and they sidestep many of the ethical concerns associated with larger vertebrate models.

The review found that glucocorticoids dominate the field. Of the 105 studies, 92 relied on these steroid drugs to trigger bone loss, with prednisolone used in 54 studies and dexamethasone in 38. The remaining studies employed a colorful menagerie of other compounds: ferric ammonium citrate to model iron overload appeared in seven studies, while doxorubicin, thioacetamide, alloxan, streptozotocin, and aluminum chloride collectively accounted for six more. Each chemical class produces bone loss through a distinct route, yet the review identified a striking convergence in the resulting pathology.

That convergence centers on a well-defined molecular cascade. Glucocorticoids enter cells and bind to cytoplasmic glucocorticoid receptors, which then translocate to the nucleus and reprogram gene expression. One consequence is the suppression of antioxidant defenses such as superoxide dismutase, glutathione, and catalase, flooding cells with mitochondrial reactive oxygen species. This oxidative stress activates NF-kappaB signaling and the RANK/RANKL/OPG axis, driving osteoclasts, the cells that demolish bone, into overdrive. Simultaneously, glucocorticoids block bone-forming pathways including focal adhesion signaling, TGF-beta/Smad, and Wnt/beta-catenin, hobbling the osteoblasts that build new bone. The result is a double hit: resorption accelerates while formation stalls, precisely the imbalance that characterizes human osteoporosis.

Iron overload models tell a parallel story. Excess iron accumulates in bone tissue, generates oxidative stress, and impairs mineralization, with studies documenting depleted levels of calcium, magnesium, potassium, and zinc in the skeleton. Doxorubicin wreaks havoc through overproduction of reactive oxygen and nitrogen species, while alloxan and streptozotocin damage osteoblasts through the buildup of advanced glycation end products. Despite these different triggers, nearly every model reproduced the same core phenotype: reduced mineralization, suppressed osteoblast differentiation, heightened osteoclast activity, and oxidative stress-mediated bone loss.

Methodologically, the field has settled into recognizable patterns. Most studies exposed larvae between 3 and 9 days post-fertilization, with prednisolone typically applied at 10 to 25 micromolar and dexamethasone at 5 to 20 micromolar. Adult fish, used in a smaller subset of studies, received higher concentrations over longer periods, sometimes up to 25 days. Assessment techniques revolve around skeletal staining: Alizarin Red to visualize calcium deposition, Calcein for real-time tracking of bone formation, and Alcian Blue for cartilage. These are complemented by quantitative polymerase chain reaction measuring osteogenic genes such as RUNX2, SP7, ALP, and OCN, alongside osteoclast markers including RANKL, CTSK, and TRAP. A handful of studies pushed the envelope further, using micro-computed tomography to measure bone mineral density or optical coherence tomography to image living bone noninvasively.

But the review also exposes a troubling weakness: methodological inconsistency. Chemical concentrations, exposure windows, developmental stages, and outcome measures vary widely across studies, making direct comparison difficult. The quality assessment was even more sobering. Only 34 of the 105 studies reported randomizing animals to groups, just 15 described random outcome assessment, and not a single study reported blinded outcome evaluation. Allocation concealment was essentially absent from the literature, and no study performed a priori sample size calculations. Many papers omitted animal numbers, ages, or routes of administration. The authors caution that researchers may have followed sound practices without reporting them, but the gap between what is done and what is documented undermines reproducibility and, ultimately, confidence in the models.

There are also inherent translational limits to keep in mind. Zebrafish lack weight-bearing bones, so their skeletons experience mechanical loads very differently from the human hip or spine. Most zebrafish studies measure mineralization and cellular activity rather than bone fragility, biomechanical strength, or fracture susceptibility, which means these models more accurately represent osteopenia-like or osteoporosis-like conditions than the full clinical disease. The review’s authors argue this should temper interpretation without diminishing the models’ genuine value for mechanistic discovery and early-stage drug screening, where zebrafish excel at rapidly identifying candidate compounds before costly validation in mammals.

To close these gaps, the review proposes a standardization framework that could reshape how the field operates. It calls for explicit reporting of developmental stage, with larvae between 3 and 9 days post-fertilization recommended for rapid mineralization screening and juvenile or adult fish reserved for structural bone assessment. Exact chemical concentrations, exposure windows, and administration routes should be specified, alongside quantitative metrics such as integrated optical density, fluorescence intensity, and vertebral counts. A core panel of molecular markers, spanning both osteogenic and osteoclastic genes, would allow results to be compared across laboratories. Transparent reporting of sample sizes, ethical approvals, and statistical methods rounds out the recommendations. If the field embraces these standards, the humble zebrafish, already swimming through hundreds of drug screens, could become a trusted bridge between cell culture and mammalian studies, accelerating the search for therapies that protect the world’s aging bones.

Subject of Research: Chemically induced osteoporosis models in zebrafish

Article Title: Experimental models of chemically induced osteoporosis in zebrafish: A systematic review

Article References: Experimental models of chemically induced osteoporosis in zebrafish: A systematic review. (n.d.). https://doi.org/10.1016/j.heliyon.2026.e45495

Image Credits: AI Generated

DOI: 10.1016/j.heliyon.2026.e45495

Keywords: zebrafish, osteoporosis, bone loss, glucocorticoids, prednisolone, dexamethasone, systematic review, bone mineralization, osteoblasts, osteoclasts, drug screening, oxidative stress