Cytokine receptor-like factor 2 B-cell acute lymphoblastic leukemia (CRLF2 B-ALL) is a high-risk subtype characterized by CRLF2 overexpression with poor survival rates in children and adults. CRLF2 and interleukin-7 receptor alpha (IL-7Rα) form a receptor for the cytokine thymic stromal lymphopoietin (TSLP), which induces JAK/STAT and PI3K/AKT/mTOR pathway signals. Previous studies from our group showed that low TSLP doses increased STAT5, AKT, and S6 phosphorylation and contributed to CRLF2 B-ALL cell survival. Here we investigated the role of TSLP in the survival and proliferation of CRLF2 B-ALL cells in vitro and in vivo. We hypothesized that high doses of TSLP increase CRLF2 signals and contribute to increased proliferation of CRLF2 B-ALL cells in vitro and in vivo. Interestingly, we observed the opposite effect. Specifically, high doses of TSLP induced apoptosis in human CRLF2 B-ALL cell lines in vitro, prevented engraftment of CRLF2 B-ALL cells, and prolonged the survival of +TSLP patient-derived-xenograft mice. Mechanistically, we showed that high doses of TSLP induced loss of its receptor and loss of CRLF2 signals in vitro. These results suggest that high doses of TSLP could be further investigated as a potential therapy for the treatment of CRLF2 B-ALL. Acute lymphoblastic leukemia (ALL) is a hematological malignancy characterized by increased proliferation of immature B or T lymphoblasts. ALL occurs in both children and adults, with a 5-year survival rate of 93% in children <15 years), 59–74% in adolescents and young adults (15–39 years), and 13–43% in adults (>40 years) 5 years, ,3,4,5.It is important to note that B-lineage ALL represents 85% of ALL cases . AdvanceCRLF2 and interleukin-7 receptor alpha (IL-7Rα) dimerize to form a heterodimeric receptor for TSLP ,16. BindThe present study was designed to investigate the role of TSLP in the survival and proliferation of CRLF2 B-ALL cells in vitro and in vivo. We hypothesized that high doses of TSLP  would increase CRLF2 signals and contribute to increased proliferation of CRLF2 B-ALL cells. Interestingly, we observed the opposite effect, with high doses of TSLP reducing viability and inducing apoptosis in human CRLF2 B-ALL cell lines in vitro. Mechanistically, we showed that high doses of TSLP-induced loss of TSLP-receptor expression and loss of CRLF2-mediated signaling in human CRLF2 B-ALL cell lines in vitro. Finally, using our novel +TSLP patient-derived-xenograft (PDX) mouse model , we showp = 0.01) and a 5.2-fold increase in Call-4 (p < 0.0001)  and MHH-Call-4  (established from 10-year-old patient at diagnosis) with increasing concentrations of TSLP  for 9 days. Cells were harvested every 3 days and stained with Annexin-V and 7-AAD to evaluate cell death using flow cytometry. We observed that high concentrations of TSLP (>200 pg/mL) decreased the viability of both cell lines ( 0.0001) C,D.p = 0.0006, 0.0025, 0.001, and <0.0001 for weeks 2, 3, 4, and 5, respectively) in leukemia burden, when compared to those treated with low doses of TSLP  to produce Mutz-5-luc cells. These cells were then transplanted into +TSLP NSG mice. Subsequent to transplanting the mice with Mutz-5-luc cells, high doses of TSLP (>200 pg/mL plasma level) were administered to the mice weekly for 5 weeks (starting 1 week after transplantation) via intraperitoneal injections (I.P.) of TSLP-producing stromal cells  in suffi of TSLP B. These  of TSLP C,D suggep = 0.0037) in the survival of the high-dose-TSLP group compared to the low-dose-TSLP group (p = 0.02) in the weight of spleens harvested from mice in the high-dose-TSLP group compared to the low-TSLP group  cells from patient-1 were used to produce a patient-derived xenograft (PDX-1). We injected TSLP-stromal cells every week  to achieve high levels (>200 pg/mL) and low levels (≤20 pg/mL) of plasma TSLP (data not shown) and followed the survival of the PDX mice. We observed a significant increase  in the xenograft mice and prolong the survival of the mice. However, these findings do not provide evidence as to whether TSLP can prevent the engraftment of leukemia cells in the mice. To address this question, we injected mice with TSLP-producing stromal cells (+T stroma) or with non-TSLP-producing stromal cells (-T stroma) weekly and evaluated bone marrow chimerism at euthanasia in the +T vs. -T groups of mice. We used CRLF2 B-ALL cells from patient-1 and patient-2 to generate the PDX-1 and PDX-2 models. Injections were started one week before leukemia cell transplantation A,B or twα + CRLF2) in CRLF2 B-ALL cells in vitro. We treated Mutz-5 and Call-4 cells with increasing concentrations of TSLP  for 24 h. Cells were harvested and stained for surface or total (surface + intracellular) expression of IL-7Rα and CRLF2, then assessed using flow cytometry or immunoblotting  proteins, which inhibit the signaling of cytokines such as TSLP via altering their pathways such as the JAK-STAT pathway . To deteThe role of TSLP as a potential therapeutic for CRLF2 B-ALL has not been previously explored. In this study we investigated whether treatment of pre-clinical models of CRLF2 B-ALL, both cell lines and PDXs, with increasing concentrations of TSLP has major effects on leukemia cell survival in vitro and leukemia burden in vivo. Our results showed that high doses of TSLP induced cell death in human CRLF2 B-ALL cell lines  via apoptosis, with higher levels observed in Call-4 compared to Mutz-5. This might be explained by Mutz-5 being more aggressive, and perhaps more therapy resistant, since they were established from a relapsed 26-year-old patient, while Call-4 cells were established from 10-year-old patient at diagnosis ,34. Anotα. A possible explanation for this is that TSLP might regulate the expression of its receptor complex by degrading the CRLF2 receptor while recycling and partly degrading IL-7Rα. Additionally, we demonstrated that high TSLP concentrations can decrease the levels of phosphorylated STAT5 and S6, suggesting an inhibition of the JAK/STAT and PI3K/AKT/mTOR pathways.Our data are consistent with a previous study that evaluated the effects of TSLP on colon cancer and reported that high doses of TSLP induced apoptosis of colon cancer cells in vitro, and decreased tumor size after peritumor injection of TSLP . The stuAnother mechanism of TSLP-mediated regulation of receptor signaling may be through SOCS proteins, which participate in the negative feedback regulation of cytokine signaling by directly interacting with JAKs or the JAK cytokine-receptor to prevent JAK phosphorylation . SOCS alOne limitation of our study is the lack of additional primary CRLF2 B-ALL samples to demonstrate the anti-leukemic effects of TSLP in a broad spectrum of patients. Additionally, delivering human TSLP (hTSLP) via injection of transduced stromal cells limits the application of this mode of delivery to human clinical trials. Despite these limitations, the current findings support the need for future studies that may incorporate a larger number of patient samples and PDXs; the evaluation of TSLP-induced CRLF2-signaling suppression as part of a multi-drug approach; and investigate a new translational method for delivering hTSLP suitable for clinical trials.To our knowledge, this is the first study showing that high doses of TSLP induce CRLF2 B-ALL cell death in vitro, reduce leukemia burden in vivo, downregulate CRLF2-receptor expression, and suppress downstream signaling. We previously showed that high doses of TSLP increased the proliferation and production of normal human CD19+ PAX+ pro-B cells and low doses of TSLP maintained the survival of CRLF2 B-ALL cells ,29. BaseHuman CRLF2 B-ALL cell lines Mutz-5 and MHH-Call-4 were obtained from DSMZ . Primary human CRLF2 B-ALL cells (patient-1 and patient-2) were obtained in accordance with protocols approved by the Loma Linda University Institutional Review Board (IRB) and with the 1975 Declaration of Helsinki, as revised in 2008. Details about patient-1 and patient-2 samples were described previously by our group . Mutz-5 For surface antigen detection, cells were stained using standard flow cytometry protocols. For total (surface and intracellular) antigen detection, cells were fixed with 1% paraformaldehyde for 10 min, then permeabilized with −20 °C-cold 100% methanol for 30 min, and then stained using standard flow cytometry protocols. To differentiate between live and dead cells, cells were stained with fixable viability dyes  as per the manufacturer’s instructions. Flow cytometry analysis was performed using a MACSQuant analyzer  and FlowJo analysis software .After 24 h of TSLP treatment, cells were rested in culture medium without cytokines for 2 h, then cultured with TSLP or no TSLP for 30 min. Cells were then harvested, fixed and permeabilized as described above for total staining, and stained for phosphorylated STAT5 and S6 proteins.TM Annexin V Apoptosis Detection Kit with 7-AAD  as per the manufacturer’s instructions.For evaluation of cell viability and cell death after treatment, cells were stained with BioLegend’s Pacific BlueEqual amounts of protein were separated with electrophoresis using SDS-PAGE . Electrophoresis was followed by protein transfer to iBlot™ 2 Transfer Stacks, PVDF, mini  using the iBlot 2 Dry Blotting System . Membranes were blocked with 5% dry milk solution prepared in TBS-T buffer  for 1 h then probed with appropriate primary antibodies, i.e., rabbit anti-CD127  or rabbit anti-CRLF2 , overnight. After at least 3 washes with TBS-T, membranes were incubated with HRP-conjugated secondary anti-rabbit IgG  for 1 h. After 3 washes with TBS-T, protein bands were detected using SuperSignal™ West Pico PLUS Chemiluminescent Substrate . Protein bands from at least 3 independent blots were scanned for each protein of interest, quantified using ImageJ software , and normalized to β-actin loaded control protein bands to determine total protein expression.All animal experiments were performed following animal protocols that were approved by the Institutional Animal Care and Use Committee of Loma Linda University. All +TSLP (high and low doses of TSLP) patient-derived-xenograft (PDX) mice were generated as described previously by Francis et al. . StromalMutz-5-luc (transduced with luciferase and with a GFP reporter) cell engraftment was monitored weekly via bioluminescent signals detected using a Lumina III IVIS instrument . A solution of 125 mg/kg D-Luciferin in PBS  was injected intraperitoneally 5 min prior to imaging. Animals were anesthetized with 1.5% isoflurane and image sequences were acquired over a period of 7 min. A series of 30 images was collected with exposures ranging from 1 to 20 s and with small (2) and medium (4) binning to ensure adequate signal sensitivity at each time point. IVIS data were analyzed using Living Image 4.5 . The #10 image  of the imaging sequence gave the best signal throughout the experiment and was selected for the time course analyses. Rectangular regions of interest (ROIs) were drawn to encompass the entire animal. Bioluminescence data are reported as total flux (p/s).Concentrations of hTSLP in sera collected from NSG mice injected with TSLP-producing stromal cells were measured using BioLegend’s LEGENDPlex™  as per the manufacturer’s instructions.Patient-1 primary human CRLF2 B-ALL cells were expanded, harvested from the bone marrow of PDX-1 mice, and cultured with 15,000 pg/mL of TSLP ex vivo for 36 h. The human CRLF2 B-ALL cells were harvested after 36 h. Human CD19+ cells were isolated via magnetic separation using the human CD19 microbead kit  as per the manufacturer’s protocol, and rapidly frozen using dry ice , CD127 PE , hTSLP-R (CRLF2) APC , hCD19 PE/Cy7 , anti-mouse CD45 FITC , PE mouse anti-Stat5 (pY694) , Cat# 612567), S6 mouse anti-(pS235/pS236) , CD10 PE , SOCS1 , SOCS3 , and IgG Alexa Fluor 647 .p < 0.05 unless stated otherwise.Statistical analyses were performed using Student’s t-test and data are expressed as mean ± SEM unless stated otherwise. Statistical significance was defined as https://patents.google.com/patent/US20210077581A1/en). Accessed on 27 December 2022. The following patent resulted from the work reported in this manuscript: US20210077581A1 (


Poor fertility is the main reason for premature culling of cattle. Dam factors and external variables can both influence how the calf fetus develops, known as fetal programming. Dam factors include age, parity, body condition, health and milk yield. External variables include nutrition and the environment during pregnancy. These all affect placental growth and the nutrient supply to the fetus, which in turn influence the size, shape and body composition of the calf. Postnatal growth rates, organ structure and immunity can all be affected. The impact on organs such as ovaries, liver, pancreas, lungs, spleen and thymus is dependent on the stage of pregnancy during which the fetal environment is sub-optimal. Regulatory systems which influence growth, metabolism and fertility can be permanently reprogrammed. Most changes affecting fertility are probably indirect. For example, calf health, the timing of puberty, age and size at first calving, and the ability to adapt to lactation after calving can all alter future reproductive potential. The size of these effects is hard to quantify due to practical difficulties in obtaining data over a sufficient time period. Nevertheless, there is compelling evidence that the fertility of some cows is compromised by events happening before their birth.Cattle fertility remains sub-optimal despite recent improvements in genetic selection. The extent to which an individual heifer fulfils her genetic potential can be influenced by fetal programming during pregnancy. This paper reviews the evidence that a dam’s age, milk yield, health, nutrition and environment during pregnancy may programme permanent structural and physiological modifications in the fetus. These can alter the morphology and body composition of the calf, postnatal growth rates, organ structure, metabolic function, endocrine function and immunity. Potentially important organs which can be affected include the ovaries, liver, pancreas, lungs, spleen and thymus. Insulin/glucose homeostasis, the somatotropic axis and the hypothalamo-pituitary-adrenal axis can all be permanently reprogrammed by the pre-natal environment. These changes may act directly at the level of the ovary to influence fertility, but most actions are indirect. For example, calf health, the timing of puberty, the age and body structure at first calving, and the ability to balance milk production with metabolic health and fertility after calving can all have an impact on reproductive potential. Definitive experiments to quantify the extent to which any of these effects do alter fertility are particularly challenging in cattle, as individual animals and their management are both very variable and lifetime fertility takes many years to assess. Nevertheless, the evidence is compelling that the fertility of some animals is compromised by events happening before they are born. Calf phenotype at birth and their conception data as a nulliparous heifer should therefore both be assessed to avoid such animals being used as herd replacements. Many previous reviews have considered factors that affect dairy cow fertility in the period before, during and shortly after breeding. The objective of this review is to go back to the time of the cow’s own conception and examine the evidence suggesting that pre-natal programming may influence her fertility as an adult. Genotype is established at fertilization and also affects fertility, but this undoubtedly important aspect is outside the scope of this article. The extent to which genetic potential is fulfilled can, however, be affected by many influences to which the animal is exposed both pre- and post-natally. The studies reported here suggest that fetal programming during pregnancy can alter size, shape and body composition, postnatal growth rates, organ structure, metabolic function and immunity. Some of these changes may act directly on the ovaries but most probably act indirectly to alter reproductive potential, implying that fertility may become compromised by events happening during pregnancy before the calf is even born.As described below, the development of the embryo and fetus are influenced by the dam’s age and size, her health and nutrition preceding and during pregnancy, fetal number and aspects of the external environment such as temperature. All of these factors and their interactions affect not only size at birth, but also how that neonate develops into adulthood. The epidemiologist David Barker first established that human babies that were born small due to poor intrauterine growth had an increased risk of developing cardiovascular disease during adult life . His worIt is now well established that perturbations to the dam’s health, nutrition and environment during pregnancy may programme permanent structural and physiological modifications in the fetus. Growth occurs throughout both embryonic and fetal development through cell proliferation together with differentiation of the individual cells into the many different cell types required to form a viable newborn animal. The destiny of each cell is influenced by signals from both its neighbouring cells and the wider environment. If the supply of either nutrients or oxygen becomes inappropriate (restricted or excess), then internal signalling mechanisms operate to alter the rates of cell division, change the pathways of differentiation or cause apoptosis ,3,4. ThiExposure of the mother to an adverse environment during pregnancy can not only influence her fetus directly, but may also have an impact on the germ cells that are forming during fetal development and may therefore also affect the fertility of the F2 generation . In humaEstablishing links between events during pregnancy and longer term postnatal development is particularly challenging in cattle. The nutritional supply to the animal itself encompasses not only the feed supplied at different stages of pregnancy, but also the dam’s body condition at the time, which affects the availability of additional nutrients through mobilisation of her body tissues ,15. StudA further complication is that most cows go through pregnancy while themselves lactating. The nutritional status of the dam is therefore impacted by her milk yield, which varies with maternal age and stage of lactation. This will in turn influence nutrient availability to the gravid uterus. Finally, the costs and practicalities of performing controlled studies and following up sufficient numbers of offspring are very considerable in cattle. Outcomes such as calf birthweight (BTW)  rather than subcutaneous fat deposition . UndernoNutritional restriction in pregnant ewes resulted in reduced growth of long bones in late gestation  and dairThe fertility of female mammals is strongly influenced by the number and quality of oocytes formed in the fetal ovary, a number which is fixed before birth . This reOvarian development in cattle fetuses begins at around 50–60 days gestation. The oocyte nests start breaking down to form primordial follicles at approximately day 80, although there is a period during which oocyte nests, primordial follicles and some developing follicles are all present . Work onThe number of antral follicles present in the ovary can be used as a proxy measurement for the size of the follicular reserve, which has previously been associated with oocyte quality . The sizImpaired intestinal development in late gestation was reported in ovine fetuses that were growth-retarded following experimental placental reduction . In contThe liver plays a central role in the regulation of metabolism and energy supply and is also critical to the working of the somatotropic axis, as it synthesizes the majority of circulating IGF-1 and IGF-2 and their binding proteins (IGFBPs) . This asThere is also evidence that fetal programming may adversely affect lung development, so making the offspring more susceptible to developing respiratory disease. Fetal lung growth is largely dependent on expansion by fluid secreted from the pulmonary epithelium . An insuBoth the thymus and spleen are major immunogenic organs for which there is evidence of fetal programming that could impair postnatal immunity. T-cell formation takes place in the thymus and lymphopoiesis occurs in the spleen, which has a key role in immunoglobulin production, lymphocyte regulation and enhancing phagocytosis ,117. A mAnogenital distance (AGD) is a feature relating to reproductive tract development that can easily be measured externally. The AGD is about twice as long in males than females and there is experimental evidence linking this measurement to pre-natal steroid hormone exposure. Treatment of late pregnant rats with adrenocorticotropic hormone (ACTH) to increase corticosteroid concentrations in their offspring resulted in an increased AGD in females at birth, but adults showed normal cyclic reproductive function . TestostInsulin produced by the pancreas is important for stimulating glucose uptake into insulin sensitive tissues such as skeletal muscle and adipose tissue. Glucose uptake by the placenta and mammary gland is not, however, regulated primarily by the insulin-sensitive glucose transporter GLUT4, so both these tissues can continue to take up glucose from the circulation in the absence of insulin stimulation ,128. AduStudies in rats and mice have shown that the intrauterine nutritional environment can cause changes in the structure and function of the fetal pancreatic islets . When ewGLUT4. Another study compared pairs of twin lambs that had at least a 25% difference in their BTWs. Both insulin and glucose tolerance were greater at 1 and 6 months in the smaller BTW lambs [In terms of postnatal effects, lambs from nutrient restricted ewes in several studies showed evidence of dysregulation in their glucose secretion, based on measurements of circulating concentrations of insulin and glucose together with intravenous glucose tolerance tests (GTT). Ford et al.  found thTW lambs . Khanal TW lambs  similarlIn cattle, Kamal et al.  undertooThe somatotropic axis, including growth hormone (GH), IGF-1 and IGF-2, and their various binding proteins and receptors, is a key regulator of growth and it also plays essential roles in many other functions including the immune and reproductive systems ,140,141.There is strong evidence that the somatotropic system in the developing fetus is modulated by prenatal programming ,79. NutrIGF1 and IGF2 mRNA were increased in the offspring of nutrient-restricted dams. Noya et al. [The fetal serum IGF-1 concentration was positively correlated to BTW in sheep  and to fa et al.  found thThese studies collectively suggest that the concentration of GH, GHR and therefore also IGF-1 during the critical first few weeks of life are strongly influenced by the pre-natal environment. These differences are likely caused by a lack of hepatic GHR in calves following undernutrition, similar to the situation experienced in postpartum cows in negative energy balance . Small nAnother endocrine system for which there is good evidence of fetal programming is the hypothalamic-pituitary-adrenal (HPA) axis ,156. A rGardner et al.  measuredLHB, FSHB or GNRH1 mRNA at either age. Another study into the effects of maternal undernutrition on the HPG axis found no effect on LH and FSH responses to gonadotropin-releasing hormone (GnRH) challenge between the female lambs derived from control or restrict fed ewes when these were tested at both 2 and 5.5 months of age. The magnitude of the pre-ovulatory gonadotrophin surge was also not affected by maternal diet [Few studies in either cattle or sheep have examined potential effects of fetal programming directly on the hypothalamic-pituitary-gonadal (HPG) axis. However, Borwick et al.  failed tnal diet . This evThe studies reported above provide strong evidence that fetal programming during pregnancy can alter organ structure, metabolic function and immunity. The follow-up periods are, however, usually quite short and few extend onwards to determine long-term effects on fertility. Apart from direct actions on the reproductive tract, many of the changes reported could potentially alter fertility through a variety of indirect mechanisms. For example, the liver and gastrointestinal tract are essential for normal metabolic function, poor lung development will increase the risk of developing respiratory disease, and the spleen and thymus are both central to immunity. If any of these malfunction in later life then cows will be more prone to developing metabolic or infectious diseases, which will in turn have a negative impact on their reproductive potential.n = 111); parities 2–3, 1.3 ± 0.05 S/C (n = 223); parities 4–5, 1.2 ± 0.06 S/C (n = 99), parity > 5, 1.5 ± 0.12 S/C (n = 60), p = 0.005) . A retrospective study by Akbarinejad et al. [The influence of dam parity on offspring fertility remains uncertain as there are limited studies and these obtained conflicting results. Swali and Wathes  found thd et al.  reportedd et al. .We also analysed the fertility data of heifers being bred for the first time according to the milk yield of their dam during her pregnancy . This shA series of experiments in sheep investigated the effects of maternal nutrition during pregnancy on aspects of reproductive performance of their lambs. Gunn et al.  comparedIn cattle, Sullivan et al.  manipulaA comparison of heifer offspring whose dams did or did not receive cooling during heat stress in late gestation found that there were no differences between the two groups for age at first service (AFS) or AFC but the cooled group of heifers required significantly fewer S/C (2.0 vs. 2.5) ,49. SurvOne potential outcome of fetal programming is low BTW. This has the advantage of being easily measurable although, as discussed above, it is not a particularly reliable guide that other important changes to the developing fetus may have occurred. Low BTW may or may not be followed by postnatal catch-up growth and it can also be associated with alterations in body proportions.One phenotype that we have observed was calves born taller at the hips than the withers (the highest point of the thoracic spine) and with a greater rump slope. This is defined as the angle between the height from the ground to the tuber coxarum and the tuber ischium . Sub-optimum growth of some heifers within each cohort was, therefore, established at an early age and resulted in animals reaching the start of breeding at an inadequate size. On the other hand, those with a greater girth measurement at 6 months or higher weight gains over the entire rearing period from 1 to 15 months required on average more S/C. These heifers presumably reached puberty earlier and were bred earlier but, although their initial conception rates were lower, they were still younger at first calving than heifers with poor growth rates [Swali et al.  found thth rates . In agreth rates ,186.n = 17 and n = 18, respectively) were too few to achieve statistical significance. Twinning is another factor leading to low BTW calves. Giving birth to twins is known to negatively impact the subsequent fertility of the mother, and heifer calves with a male sibling become infertile freemartins [Perhaps surprisingly, Swali and Wathes  found themartins . There demartins .Another study investigated the influence of bodyweight at first breeding in beef heifers on the reproductive performance of their offspring . Angus-cAnogenital distance is an accepted biomarker for prenatal androgenization and fertility in humans and rodents, so a series of studies have investigated its potential link to cattle fertility. Initial work in Canadian Holstein cows found that AGD was highly variable with a normal distribution, and the measurement was weakly associated with age and height. There was an inverse relationship between AGD and conception data in first- and second-parity cows but this was not evident in older animals . NullipaEvidence presented above suggests that undernutrition in late fetal life can influence pancreatic insulin secretion directly and also cause differential insulin sensitivity between tissues, which will in turn alter their prioritization for glucose utilization. This will affect both the amount of adipose tissue deposition and the supply of energy going into the mammary gland for milk production. Van Eetvelde and Opsomer  reviewedBrickell et al.  found thThere is a well established relationship between circulating concentration of IGF-1 in the early postpartum period and cow fertility e.g., ,202,203),203202,2A large body of work has investigated the long-term effects of dam morbidity during pregnancy on her offspring. This has mainly focussed on a wide variety of infectious diseases  which can cause placentitis and/or be transmitted to the developing fetus, causing abortion, developmental abnormalities or postnatal infection . Other iThe evidence reviewed above also indicates that fetal programming caused by suboptimal dam nutrition can affect the development of the thymus, spleen and lungs, all of which might be expected to put calves at greater risk of catching enteric and bovine respiratory disease (BRD), particularly during the pre-weaning period. Some studies have therefore investigated the relationship between calf BTW and disease risk. Corah et al.  and WindOverall, the relationships between BTW and calfhood disease are not very strong. One possible explanation for this could be that the critical windows of development which most affect the intestines, lungs and immune system do not necessarily result in measurable differences in BTW. Much stronger associations have, however, been found between morbidity and circulating IGF-1. Johnson et al.  showed tAlthough relatively little is known about this topic, it deserves a mention in the context of cattle fertility. Epigenetic modifications have been detected in bull sperm, which gave rise to preimplantation embryos with significant differences in their transcriptome (reviewed by ). This pAnother topic not already covered is the widespread use in cattle of assisted reproductive technologies (ART), including AI and embryo transfer (ET). In humans, there is evidence that ART can affect infant BTW and glucose metabolism, effects attributed to epigenetic changes in the early conceptus . This isp = 0.06); AFC 698 v 663 days, p = 0.1). These results need to be interpreted with caution as different bulls were used between groups and there were only 38–43 heifers per group. Differences recorded around birth  could also have been influential in this study. Another larger retrospective analysis compared the fertility of offspring arising from in vitro embryo production  with offspring from multiple-ovulation embryo transfer . These techniques were both performed in the same facility over a 10-year period using animals balanced for genetic merit for breeding values. The IVP embryos were subsequently slightly more fertile as heifers than the MOET group, needing 1.9 vs. 2.0 straws to become pregnant, and with a 56 day nonreturn rate of 61.7% vs. 59.6% [With respect to fertility in cattle, one study compared the fertility of heifer offspring produced by AI or ET using either fresh (ET-F) or vitrified (ET-V) in vitro produced embryos . No diffs. 59.6% . A simils. 59.6% . There ws. 59.6% .Efforts by breeders to reverse negative trends in cow fertility caused by the previous overemphasis on production traits have led to small improvements in recent years. This follows greater inclusion of fertility and survival traits in genetic selection indices ,233 and Nevertheless, some generalisations can be made. In practice, calves that are more likely to have problems with fertility can be identified before they are bred based on their appearance, size measurements (including height as well as weight) and growth rates. Taking a side view photograph of each pre-weaned heifer in her first week of life is a quick and simple action in dairy herds and this can be stored alongside other records of growth rates and health. This information should then be considered along with genotype when deciding which animals to use as herd replacements so that heifers with a poor phenotype can be avoided. Rearing unsuitable heifers to breeding age carries a significant cost as these animals are unlikely to ever become profitable . Not allA greater awareness is also needed that any perturbations which affected the pregnant dams could have a permanent adverse effect on their developing calves. These could include a period of inappropriate nutrition (under- or over-feeding), adverse weather, or an episode of infectious or metabolic disease. Their impact will, however, be modified by factors such as their duration, stage of pregnancy, age of dam, milk yield, body condition etc, all of which will interact with each other. For example, NP heifers are likely to be the ones most greatly influenced by a period of either under- or over-nutrition during pregnancy. These factors and some of their possible interactions are illustrated in


In 2021 a new INSTI, cabotegravir, was approved by the FDA for HIV-1 treatment. We thus set out to evaluate its efficacy against HTLV-1 infection in vitro. Strand transfer assays performed using recombinant HTLV-1 integrase treated with increasing concentrations of cabotegravir, effectively inhibited strand transfer activity, displaying an IC50 of 77.8 ± 22.4 nM. Furthermore, cabotegravir blocked HTLV-1 transmission in tissue culture; we determined an EC50 of 0.56 ± 0.26 nM, similar to bictegravir. Alu-PCR confirmed the block in integration. Thus, there are four INSTIs and one reverse transcriptase inhibitor approved by the FDA for HIV-1 treatment, that potently block HTLV-1 infection in vitro. This should strongly encourage the establishment of a new standard of HTLV-1 treatment – particularly for pre-exposure prophylaxis and prevention of mother-to-child transmission.Human T-cell lymphotropic virus type 1 (HTLV-1) is a deltaretrovirus most prevalent in southwestern Japan, sub-Saharan Africa, Australia, South America, and the Caribbean. Latest figures approximate 10 million people worldwide to be infected with HTLV-1. This is likely a significant underestimation due to lack of screening in endemic areas and absence of seroconversion symptoms. The two primary diseases associated with HTLV-1 infection are adult T cell leukaemia-lymphoma, a malignant and, sometimes, aggressive cancer; and HTLV-1 associated myelopathy/tropical spastic paraparesis, a debilitating neurological degenerative disease. Unfortunately, despite the poor prognosis, there is currently no effective treatment for HTLV-1 infection. We previously showed that integrase strand transfer inhibitors (INSTIs) clinically used for human immunodeficiency virus type 1 (HIV-1) prophylaxis and treatment are also effective against HTLV-1 transmission  Three decades ago, infection with human immunodeficiency virus type 1 (HIV-1), a close relative of human T-cell lymphotropic virus type 1 (HTLV-1), was almost universally fatal. Introduction of antiretroviral therapy with drugs targeting the viral reverse transcriptase (RT) enzyme and protease enzymes, seropositive patients got a second chance to live. Since 2007, a novel class of inhibitors, targeting the viral integrase (IN) enzyme, integrase strand transfer inhibitors (INSTIs) completed the current list of approved antiretrovirals . GeneratHuman T-cell lymphotropic virus type 1, discovered shortly before HIV-1 , infectsDeveloping disease is positively correlated with the proviral load (PVL) (≥4%), and in the case for ATLL, an oligoclonality index determined by flow cytometry >0.77 increases the risk of developing disease . Use of in vitro    in the lin vitro strand-transfer activity was measured by means of a strand-transfer assay. A schematic illustrating the assay is shown in 2, 5.8 μM ZnCl2, 12.8 mM DTT, 0.53 μM donor vDNA, 1.6 μM HTLV-1 IN. This was then incubated in the presence of a given concentration of CAB (dissolved in DMSO) or with DMSO alone  for 30 min at room temperature. A total of 300 ng of target DNA [supercoiled pGEM-9Zf(-)] was then added to initialise the reaction, which was carried out at 37°C for 30 min. Once completed, samples were deproteinised and DNA products precipitated as described previously (Human T-cell lymphotropic virus type 1 integrase (IN) was purified as reported previously . Inhibiteviously , 18. Reswhere SE denotes the standard error computed by Prism.2. HTLV-1 infection was conducted as previously outlined  and the target cell line (Jurkat cells) were co-cultured in serum free medium, in the presence of increasing concentrations of CAB and 15 mM MgCl2. This resulted in a culture of 0.5e6 of each cell type in a total of 500 μL. Following a 17.5-h infection, cells were washed with PBS and mixed with 25 μL magnetic anti-CD25 beads  to enable depletion of MT-2 cells from the culture (The MT-2 and Jurkat (E6.1) T cell lines (ATCC) were maintained in Roswell Park Memorial Institute (RPMI) media supplemented with 10% heat inactivated foetal bovine serum (Gibco), 100 U/mL penicillin, 100 μg/mL streptomycin, and 0.25 μg/mL fungizone (Gibco) (referred to as complete medium). Cells were cultured in a humidified atmosphere at 37°C with 5% COoutlined , 17. In  culture , 17. Bot culture . Cell vitax gene and human ALBUMIN. Copy numbers of each gene were quantified via comparison to standard curves generated from clone 11.50 , or uninfected control, respectively. With the knowledge that there is one copy of tax gene and two copies of ALBUMIN per cell, PVL was calculated [(2 × tax)/albumin]. Data was normalised to DMSO, which was set to 100%. Dose-response curve fitting was done in Prism 8. Cumulative standard deviations were calculated as described above for the in vitro IC50 data. The 200 nM and DMSO samples underwent Alu-qPCR to quantify integrated DNA, as previously explained . BIC, however, is much more potent in cell culture  was achieved through combination therapy with zidovudine (RT-inhibitor) and the HDAC inhibitor valproic acid . In huma50 = 17.8 ± 7.2 nM)  . TherefoJapan is the only country where pregnant women are screened for HTLV-1, and infected mothers are closely monitored and strongly advised to formula feed their babies . AlthougFrom HIV-1 research, we know that both the pregnant woman and the developing embryo are safe when the mother uses RAL or TDF during pregnancy , 48. We 1 following two large studies which illustrated that bimonthly CAB injections reduced the risk of HIV transmission more than daily PrEP pills. Given that >70% of HTLV-1 transmission occurs via condomless intravaginal sex, and the knowledge that INSTIs and TDF efficiently block HTLV-1 transmission in tissue culture, it is time to consider PrEP to prevent HTLV-1 transmission, and possibly prioritise CAB-based PrEP in HTLV-1 endemic settings.In December last year, the FDA approved the use of long-acting injectable CAB for HIV PrEPFinally, whilst we urgently need to know if we can effectively block HTLV-1 sexual transmission and MTCT by using INSTIs with or without TDF, we should not forget the people who inject drugs, and patients who depend on organ donor transplantations – they too could profit from PrEP.The original contributions presented in the study are included in the article/in vitro strand transfer assays. BS did the HTLV-1 transmission experiments. GM designed and led the research. GM, BS, and MB wrote the manuscript. All authors contributed to the article and approved the submitted version.MB performed the The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Psoriasis is a chronic papulosquamous skin disease with an autoimmune pathogenic traits and strong genetic predisposition. In the past few decades, with the rapid development of molecular biology and cell biology, the inherent pathogenesis of psoriasis has been gradually elucidated, in which cytokine inflammatory loops, cell signaling pathways, and epigenetic factors such as miRNAs have been demonstrated to play important roles in regulating the development and progression of psoriasis. More importantly, understanding the pathogenesis of psoriasis has promoted the development of effective treatment for psoriasis. In this review, we systemically summarized the molecular mechanisms regulating the development and progression psoriasis, introduced various therapeutics used for clinical psoriasis therapy, and highlighted the recent advances in nanoparticles (NPs)-mediated drug delivery for psoriasis treatment. In addition, emerging evidences have revealed that epigenetics especially microRNAs (miRNAs)-mediated target gene expression plays an important role in the development and progression of psoriasis.Psoriasis is an immune-mediated and chronic-hereditary inflammatory skin disease, which affects around 2% of the global population and imposes a heavy psychological and physical burden on patients –3. The petc.) –7. Multivia the administration of vitamin D3 or glucocorticoids complemented with phototherapy. For the patients with advanced psoriasis, systemic therapy is the main therapeutic modality, which usually involves the use of methotrexate (MTX), cyclosporine A, or biotherapeutics targeting cytokines   –10. In tkinumab) . TherefoOver the past few decades, nanoparticles (NPs)-mediated drug delivery has been widely used for the treatment of various diseases, showing the advantages of promoting drug water solubility, enhancing drug stability and bioavailability, reducing toxic and side effects, and improving therapeutic outcomes . Inspiree.g., IFN-γ, TNF-α, and IL-1β) have been found to play an important role in regulating the development and progression of psoriasis  is an important signaling hub that coordinates cellular and tissue responses. The upstream pathways of mTOR include PI3K-AKT and LKB1/CD73-AMPK. The PI3K/AKT/mTOR signaling pathway is widely involved in the proliferation of various types of cells, and also plays an important role in regulating development and progression of autoimmune diseases including psoriasis Figure 2i.e., JAK1, JAK2, JAK3, and tyrosine kinase 2 (TYK2). The STAT family includes seven subtypes    is an intracellular signaling pathway that enables extracellular signals to be transmitted to the nucleus through relatively few intermediate links, thereby enabling rapid cellular responses to various signals reaction Figure 3d STAT6) . Many did TNF-α) . Therefod TNF-α) . In factd TNF-α) .e.g, IL-6, IL-1β, and TNF-α) to accelerate the progression of psoriasis (Mitogen-activated protein kinase (MAPK) is a member of the serine-threonine kinases kinase family. Its signal is transmitted from the cell membrane to the nucleus and is one of the most important signaling pathways that control cells proliferation Figure 4soriasis , 47. NF-soriasis . At presBesides the signaling pathways described above, some other canonical signaling pathways have been also reported to involve the pathogenesis of psoriasis. For example, as a highly conserved signal transduction pathway that coordinates the regulation of epidermal cell proliferation, differentiation, and migration, dysregulation of Notch signaling pathway is closely associated with various human skin diseases including psoriasis –51. In aWith the deepening of psoriasis-related pathogenesis research, some potential psoriasis-related signaling pathways have also been discovered. For example, Müller A et al. demonstrated that cyclin-dependent kinase 4 (CDK4) and CDK6 can induce the activation of STAT3 after phosphorylation of methyltransferase (EZH2) in keratinocytes, thereby inducing the production of IκBζ (a key pro-inflammatory transcription factor required for psoriatic cytokine synthesis) . Yan et e.g., proliferation, differentiation, and apoptosis) (MicroRNAs (miRNAs) are small and highly conserved non-coding RNA with a length of about 22 nucleotides, which can bind to specific target mRNA and play a negative regulatory role by promoting the degradation of target mRNA and inhibiting translation, and participates in biological process (optosis) , 58. In optosis) , 60. So via regulating the STAT3 and SAPK/JNK signaling pathways  are used to target TNF-α signaling pathway  and psoriasis therapy.In the past two decades, the application of NPs for drug delivery has introduced exciting opportunities to improve the therapeutic outcomes of various diseases including psoriasis, showing the advantages of improving the drug stability, improving the drug accumulation in the disease sites, and reducing the toxic and side effects. Inspired by these unique advantages, numerous NPs have been designed and developed for the delivery of various types of therapeutics drugs  after topical administration to an IMQ-induced psoriatic plaque model  is one of the great technologies discovered in the biological field. Currently, CRISPR-Cas9 technique has become one of the most powerful tools for disease treatment . The mec30% NPs) with an average size of less than 15 nm, which could penetrate the stratum corneum and preferentially enter keratinocytes when applied to a mouse model of IMQ-induced psoriasis. More importantly, these modified Au NPs could reduce the pro-inflammatory response induced by the IL-17 signaling pathway and show the similar therapeutic efficacy as standard steroids and vitamin D analogs without causing hair loss and skin wrinkles  in psoriasis by blocking of the NF-κB signaling pathway in macrophages  and gold nanoparticles (Au NPs) have been developed for the treatment of psoriasis with powerful anti-inflammatory effect –86. For rophages . Besidessoriasis .via impairing the secretion of inflammatory cytokines such as TNF-α and IL-23 (Transdermal drug delivery is an effective route for the treatment of skin diseases, because it allows direct targeting of lesions on the skin and reduces the side effects associated with systemic drug delivery –93. Howend IL-23 .Although the pathogenesis of psoriasis is diverse and complex, cytokine inflammatory loops, cell signaling pathways, and epigenetic factors such as miRNAs have been widely recognized as the important factors regulating the development and progression of psoriasis. Based on these regulatory mechanisms, a variety of therapeutic drugs including small molecule drugs and biologics have been designed and approved for clinical treatment of psoriasis. Although these drugs have achieved great success in clinic, much more efforts need to be paid to address their drawbacks, especially the toxic and side effects. The use of NPs for drug delivery has provided the valuable chances to promote the therapeutic outcomes of various types of drugs and thus provided innovative insights into the treatment of psoriasis. However, the clinical translation of these nanodrugs is still challenged and more efforts are required to promote their clinical use, especially needing to solve the difficulty in controllable and reproducible synthesis of nanodrugs and their scalable manufacturing. With rapid development of molecular biology and nanomedicine, we believe that more types of therapeutics will be definitely developed and marked for the treatment of psoriasis.QS, RLiu, ST, and JF wrote the manuscript. QS, XX, JF, and RLi revised the manuscript. All authors contributed to the article and approved the submitted version.


Premature ovarian insufficiency (POI) is a female reproductive disorder of unknown etiology with no definite pathogenesis. Melatonin (MT) is an endogenous hormone synthesized mainly by pineal cells and has strong endogenous effects in regulating ovarian function. To systematically explore the pharmacological mechanism of MT on POI therapy, a literature review approach was conducted at the signaling pathways level.  Relevant literatures were searched and downloaded from databases, including PubMed and China National Knowledge Infrastructure, using the keywords “premature ovarian insufficiency,” “Hippo signaling pathways,” and “melatonin.” The search criteria were from 2010 to 2022. Text mining was also performed.  MT is involved in the regulation of Hippo signaling pathway in a variety of modes and has been correlated with ovarian function.  The purpose of this review is to summarize the research progress of Hippo signaling pathways and significance of MT in POI, the potential crosstalk between MT and Hippo signaling pathways, and the prospective therapy. Ovarian aging is a complex physiological process with multiple factors interacting and gradually accumulating, the essence of which is the decrease in the number and quality of follicles in the ovary, which is reflected in the reduction of reproductive and endocrine functions until they are lost . HoweverThe Hippo signaling pathway is currently a hot topic among researchers. Studies have shown that the Hippo/Yes-associated protein (YAP) signaling pathway can regulate ovarian cell aging. For example, human umbilical cord mesenchymal stem cell-derived exosomes improve ovarian function and proliferation of POI by regulating the Hippo signaling pathway . Even thMelatonin (MT), commonly known as a sleep hormone, controls physiological processes such as regulation of sleep-wake rhythms, body temperature, and physiological activity in circadian rhythms. MT also regulates ovarian function and is associated with the Hippo signaling pathway . FurtherDespite recent advances in the field of reproductive endocrinology, the pathogenesis of POI remains unclear, which limits the development of POI treatment and has become a hot spot for researchers to gather , 11. WhaFor various reasons, POI is a condition in which ovarian failure occurs after menarche and before the age of 40 due to the destruction or depletion of follicles in the ovaries, causing a range of symptoms such as menstrual disorders and loss of libido . POI canThe incidence of POI in the general population of women is approximately 1-2% and a Swedish survey shows an increasing trend in the prevalence of POI in recent years . The larPatients with POI can suffer from a range of symptoms due to low estrogen, such as menstrual cycle disorders or even amenorrhea; hot flashes; and night sweats, irritability, and low libido, as well as symptoms of perimenopausal syndrome such as osteoporosis, and reduced fertility or even infertility which happened to some patients , 21. AlsFollicles are the structural units of the ovary, comprising a single oocyte surrounded by supporting somatic cells. Folliculogenesis starts in fetal life with primordial follicle development, the majority of which will remain dormant. Folliculogenesis is a complex phenomenon that starts with the initial recruitment and activation of a primordial follicle. After activation, primordial follicles grow and mature into primary, secondary, and finally antral follicles. The mechanism underlying the developmental progression of human primordial follicles is unclear. However, the initial recruitment has been linked to protein kinase B (Akt) and mammalian target of rapamycin (mTOR) signaling pathways , 22.Folliculogenesis and ovulation in normal ovary differ from that in POI ovary (impaired folliculogenesis). Under the regulation of intraovarian factors and gonadotropins, primary follicles develop into pre-antral and early antral follicles, which are the most susceptible to atresia, or follicle death. Then, they become preovulatory follicles, resulting in oocyte release and corpora lutea formation. Defects in folliculogenesis  cause POI . Most ofAnti-Müllerian hormone (AMH) belongs to transforming growth factor beta superfamily. AMH expressed by pre-antral and small follicles granolosa cell is suggested by recent studies the best biomarker of ovarian reserve currently available . With inThe detectability of serum AMH in POI patients could be remarkably related to the presence of 15 or more follicles in their ovaries , 31, 32.α are the major cytokines involved in this process [The main mechanisms of POI pathogenesis like oxidative stress and inflammation are related to phosphatidylinositol 3-kinase (PI3K)/Akt pathway. Necrosis and necroptosis are involved in germ cell depletion from the mammalian ovarian cohort . Oxidati process , 36.In the PI3K/Akt signaling pathway, once Akt is activated, it causes further cascades in the signaling pathway. Phosphorylated Akt phosphorylates a range of downstream target proteins, including forkhead box O 3 , Bcl-2-associated death promoter , mTOR (controls protein biosynthesis and regulates cell growth), and p27 (maintains primary follicular stores) –39. HowePOI diagnosis and treatment guidelines suggest hormone replacement therapy (HRT) as the main treatment, as well as stem cell therapy and traditional Chinese medicine . ComprehMT can delay ovarian aging, regulate ovarian biorhythm, promote follicle formation, and improve oocyte quality and fertilization rate . As a frWhat's more, MT can act on the hypothalamic-pituitary-ovarian axis (HPO) via hypothalamic gonadotropins or directly bind to ovarian granulosa cells to exert effects on HPO . AdditioIt has been found that long-term application of MT can slow down ovarian aging. MT delays ovarian aging through a variety of mechanisms, including antioxidation, DNA repair, telomere maintenance, silent information regulator (SIRT) family activity, ribosomal function, and autophagy , 63. SIRIn the ovary, oocytes and somatic cells produce reactive oxygen species (ROS) and reactive nitrogen species (RNS) in the follicular microenvironment. These oxidants regulate the molecular and biochemical pathways involved in follicle formation, thereby destroying oocytes and causing follicular atresia , 60. OxiA growing body of data suggests that MT mediates reproduction by interacting with MT1 and MT2 in the ovary , 70. In The ovarian reserve is constituted by the quality and quantity of the primordial follicles, which both decline with increasing age . The numThe expression of silent information regulator family  in the ovary is positively correlated with ovarian reserve . These pThe biological clock system plays an essential role in the physiological activity of the ovary and is involved in the regulation of ovulation, steroid hormone synthesis, and oocyte maturation . Disturbα subunit and Gβγ complex interact with various effector molecules involved in cellular signaling [As mentioned above, MT can signal through both MT1 and MT2 G-protein coupled receptors (GPCR). Activation of the receptors causes dissociation of the heterotrimeric G-proteins and the resulting Gignaling . Recentlignaling   through subsequent activation of protein kinase A (PKA) and protein kinase C (PKC) [αs-mediated activation of PKA and PKC in response to different stimuli  may inhibit cell proliferation and invasion through a series of convergent mechanisms, including large tumor suppressor 1 and 2 (LATS1/2) activation, inhibition of nuclear factor-κB (NF-κB) transcriptional activity, and suppression of the androgen receptor (AR) response in AR-positive cells [κB, suggesting that MT may potentially inhibit YAP/TAZ oncogenic function by increasing LATS1/2 activity (following PKA and PKC activation) or reducing TAZ transcription (following NF-κB inhibition) [YAP and transcriptional co-activators with TAZ are major targets of the Hippo signaling pathway and play important roles in the regulation of development, homeostasis, and regeneration –93. GPCR C (PKC) –97. Thusve cells , 98, 99.ibition) .It has been shown that the Hippo signaling pathway is closely related to the tissue metabolic pathway and they work together to regulate cell proliferation, differentiation, and apoptosis . GlucoseIn addition to GPCR signaling, YAP/TAZ is regulated by intercellular contact, mechanical force, and metabolism. They affect YAP/TAZ function by inducing specific intracellular signaling through Hippo kinase cascade-dependent and non-dependent mechanisms. Some of these mechanisms may be cross-linked with MT signaling. In conclusion, these evidences indicate that there may be crosstalk between MT signal, Hippo signal, and insulin-glucagon signal  Figure .Sirtuins, a family of NAD+-dependent deacetylases, have recently emerged as key metabolic sensors of body homeostasis. Together, they respond to metabolism, inflammatory signals, or oxidative stress, and have been linked to aging and longevity , 105. ThStudies based on the SIRT1 activator SIRT1720 or resveratrol support the idea that targeting SIRT1 as a main factor in the regulation of follicular dynamics may be a promising strategy for the prevention of ovarian aging , 107. ThSIRT1 coordinates the adaptive response of mouse oocytes to oxidative stress, possibly by promoting the activity of FOXO3 and SOD2 . The SIRβ toxicity. It also activates processes in an anti-inflammatory network, in which SIRT1 activation, upregulation of nuclear factor erythroid-2-related factor 2 (Nrf2), down-regulation of NF-κB, and release of the anti-inflammatory cytokines IL-4 and IL-10 are involved [Aging and various age-related diseases are associated with reduced MT secretion, pro-inflammatory changes in the immune system, and reduced SIRT1 activity –114. MT,involved , 119.The FOXO family of transcription factors  regulates the expression of genes associated with inhibition of cell cycle progression, hematopoietic differentiation, and resistance to stress . FOXO3 hBonni demonstrated that the core factor Mst1 in the Hippo pathway can phosphorylate FOXO3 (followed by FOXO1), mainly Ser207 (Ser212 in FOXO1), which is a conserved site in the forkhead domain . This phMany molecules in the Hippo signaling pathway are in connection with the regulation of oocyte growth, primordial follicle development, and granulosa cell proliferation and differentiation . AbnormaApart from follicular activation, the Hippo signaling pathway in POI has been barely studied, with only a few reports suggesting that it may be part of the mechanism of action of herbal or natural products. For example, Xie and colleagues demonstrated that the Huyang yangkun formula enhanced ovarian function in POI rats by repairing the dysfunction of the Hippo-JAK2/STAT3 signaling pathway . Ai and The Hippo signaling pathway in granulosa cells plays an important role in regulating the proliferation and differentiation of granulosa cells induced by gonadotropin and acts as crucial factor in the growth and maturation of activated follicles . KnockdoFor Hippo pathway genes, LATS1 deficiency in mice causes sterility and ovarian tumourigenesis, while LATS1 regulates the activity of FOXL2, which is a defective gene in some POI patients , 136.The application of Hippo-specific inhibitors can provide strong evidence for the study of the correlation between follicular development and Hippo signaling pathway and provide guidance for the development of POI biological gene therapy or anti-POI drugs. However, the number and structure types of Hippo inhibitors reported are quite limited, and few clinical studies on the treatment of POI are conducted.The pathogenesis of POI is complex and regulated by a combination of multiple factors and components. Numbers of studies on mammalian MT and Hippo signaling pathways have made progress in understanding the MT and Hippo action mechanism on POI . There a


Retroviridae and share the same tropism, e.g., human CD4+ T cells. These viruses show significant similarities in the viral genomic structure and the molecular mechanism of the replication cycle. However, HTLV-1 and HIV-1 infected T cells show different phenotypes, especially in the level of virion production. We speculate that how the activity of HTLV-1 Rex and its counterpart HIV-1 Rev are regulated may be closely related to the properties of respective infected T cells. In this review, we compare various pathological aspects of HTLV-1 and HIV-1. In particular, we investigated the presence or absence of a virally encoded “regulatory valve” for HTLV-1 Rex or HIV-1 Rev to explore its importance in the regulation of viral particle production in infected T cells. Finally, wereaffirm Rex as the key conductor for viral replication and viral pathogenesis based on our recent study on the novel functional aspects of Rex. Since the activity of Rex is closely related to the viral replication rate, we hypothesize that the “regulatory valve” on the Rex activity may have been selectively evolved to achieve the “scenario” with early viral particle production and the subsequent long, stable deep latency in HTLV-1 infected cells.HTLV-1 is an oncovirus causing ATL and other inflammatory diseases such as HAM/TSP and HU in about 5% of infected individuals. It is also known that HTLV-1-infected cells maintain a disease-free, immortalized, latent state throughout the lifetimes of about 95% of infected individuals. We believe that the stable maintenance of disease-free infected cells in the carrier is an intrinsic characteristic of HTLV-1 that has been acquired during its evolution in the human life cycle. We speculate that the pathogenesis of the virus is ruled by the orchestrated functions of viral proteins. In particular, the regulation of Rex, the conductor of viral replication rate, is expected to be closely related to the viral program in the early active viral replication followed by the stable latency in HTLV-1 infected T cells. HTLV-1 and HIV-1 belong to the family  Subsequently, it spread throughout the world with the geographic movement of people. Currently, seven types of HTLV-1a-g have been identified, and HTLV-1a has an additional eight subtypes, A-F and Senegalese. There is little genomic variation among the HTLV-1 types, indicating that the virus has spread to the human world ears ago . Among sears ago , 14, whiears ago . Thus, Hin vitro infection experiments and in vivo animal model  and that 77% of cases are transmitted from men to women . The avein vitro and in vivo  by Tax for HTLV-1 (via a cellular transcription factor CREB (cAMP response element binding protein). The Tax-CREB complex then recruits strong cellular co-activators such as p300 (E1A binding protein p300), CBP (CREB binding protein), and PCAF (P300/CBP-associated factor) to activate the LTR  in the 3′ LTR  and/or the virus-specific system  to optimize the viral gene expression level beneficial for replication.From the provirus of HTLV-1 and HIV-1, the doubly spliced mRNAs encoding accessory proteins, the singly spliced mRNA encoding Env, and the unspliced mRNA encoding Gag/Pro/Pol are transcribed from the provirus. At first, depending on the host cellular pathway, the doubly spliced e 3′ LTR Figure 2e 3′ LTR , 76. HTLr export , 57, 77.r export –80. NXF1r export . Therefor export . Indeed,r export , probablr export . Each covia multimerization of Rev , but not the 3′ LTR, of provirus are highly methylated , 118. ReThe proviral methylation has been also reported in HIV-1 infected cells , 122. Itde novo infection are proposed by in vitro/in vivo experiments  is kept under 4% in most asymptomatic carriers  in an infected individual is reduced to the set point, yet never be undetectable level without treatment even during chronic infection  Figure, 126 F, thus sovia export and stabilization of unspliced/singly spliced viral structural mRNAs  of these viruses appear to be the regulation and reactivation of viral production in the host cell. As explained above, Tax and Rex for HTLV-1 and Tat and Rev for HIV-1 are essential for viral replication . Particual mRNAs . TherefoTax/Rex mRNAs are RxRE/CRM1-dependently transported out of the nucleus  RNA, which is the only viral factor to suppress the viral gene expression from 5′LTR (as L.545) via epigenetic modification and replication, thus asp mRNA is considered to play an important role in the maintenance of the latent infection of HIV-1  LTR of HIV-1 provirus is not methylated universally, a leaky viral expression continues at a low level in latency. No regulatory valve on the Rev pipeline and leaky suppression of viral gene expression may be related to the shallow latency of HIV-1 infected cells, which are easily reactivated  How does Rex avoid unnecessary splicing?, 2) Why does Rex preferentially use CRM1, a major nuclear export adapter of the host cell?, and 3) Does the RxRE/CRM1 dependent pathway contribute to the selective translation of viral RNAs? We speculate that Rex has as yet unknown aspects to escort viral RNAs from the transcription to the translation. To explore new aspects of Rex, we recently conducted transcriptome and interactome analysis of Rex . Firstlypathways . Recentlpathways . Indeed,pathways Figure 5via the transport of viral RNA. In this review, we note that HTLV-1- and HIV-1-infected T cells have different characteristics, despite the many similarities in the functional mechanisms of these accessory proteins. By exploring the similarities and differences between HTLV-1 and HIV-1, we proposed a novel possibility that the presence or absence of a virus-derived regulatory valve for Rex or Rev activity, respectively, may relate to the permissiveness of the viral gene expression, thus the phenotype and pathogenesis of the infected cell. Our recent study, which demonstrated the involvement of HTLV-1 Rex in a much wider range of host-cellular pathways than ever expected  and (19K07573) to KN, and (20KK0181) to TW.We thank very much all the corroborators and students in the Department of Computational Biology and Medical Scienced, Graduate School of Frontier Sciences, The University of Tokyo, for their participation and assistance in the research of HTLV-1 Rex.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


A significant body of research has demonstrated that antidepressants regulate neurotrophic factors and that neurotrophins themselves are capable of independently producing antidepressant-like effects. While brain derived neurotrophic factor (BDNF) remains the best studied molecule in this context, there are several structurally diverse trophic factors that have shown comparable behavioral effects, including basic fibroblast growth factor (FGF-2), insulin-like growth factor 1 (IGF-1) and vascular endothelial growth factor (VEGF). In this review we discuss the structural and biochemical signaling aspects of these neurotrophic factors with antidepressant activity. We also include a discussion on a cytokine molecule erythropoietin (EPO), widely known and prescribed as a hormone to treat anemia but has recently been shown to function as a neurotrophic factor in the central nervous system (CNS). Neurotrophic factors are a family of nervous system proteins that play essential roles in neuronal development, survival, plasticity and function. The neurotrophic factor hypothesis of depression and antidepressant action continues to be highly influential in the field of biological psychiatry, providing a firm molecular neurobiology framework for the behavioral actions of antidepressants. Over the years since its inception, supportive evidence has steadily accrued. Neurotrophic factors have been implicated in the molecular and behavioral actions of multiple classes of antidepressants, including selective serotonin reuptake inhibitors , neurotrophin 3 (NT3) and neurotrophin 4 (NT4). Since the report of hippocampal BDNF and Tropomyosin receptor kinase B (TrkB) mRNA induction by electroconvulsive seizure and multiple classes of chronic chemical antidepressants  than BDNF  family has 18 ligands that signal through four known tyrosine kinase receptors, fibroblast growth factor receptors, FGFR1–FGFR4  anemia drug , a chemically engineered non-erythropoietic derivative of EPO, was therefore a significant advancement as it dissociated the neurotrophic activity of EPO from the erythropoietic effects  TheIn this regard, it is important to note that it is the contribution of trophic signaling to neuronal structure and function in key brain circuits that eventually produces changes in behavior. The cellular phenomena that they regulate such as neurogenesis, gliogenesis and synaptogenesis are important determining factors as to whether the effects are long lasting or short term. There are however questions that can only be answered by further research. As shown in multiple instances, blockade or inhibition of trophic signaling  results in a loss of the antidepressant effects of conventional antidepressants. Why does the inhibition of one pathway truncate behavioral effects? Perhaps this is due to crucial common targets downstream trophic cascades that are subject to control upstream. Can inhibition of trophic signaling in non-depressed individuals lead to depression? Several tyrosine kinase inhibitors are in use to block growth factor signaling in cancer , grant MH106640 (SN) and the University of South Dakota Center for Brain and Behavioral Research. The funding agency had no role in writing of the manuscript.


Introduction: Depression is an emotional disorder associated with morbidities and disabilities worldwide. The growing use of antidepressants is a concern for health managers because there are still unanswered questions on the effectiveness and safety of these medicines. Drug sales have increased in Brazil in recent years, but investigations on antidepressants sales are not available. We aimed to describe the trends in the antidepressant commerce in Brazil in a five-year period.Materials and Methods: We performed an ecological study on antidepressant sales in Brazil, from November 2014 to October 2019, using data from IQVIA™, a data provider of pharmaceutical sales. Antidepressants were coded by the Anatomical Therapeutic Chemical classification system, and sales were presented in defined daily doses (DDDs) and DDDs per 1,000 inhabitants per day (DIDs). The results were expressed in absolute quantities and growth rates.Results: The analyzed dataset contained 23 active substances in 780 products. The total sales of antidepressants increased from 23.3 DIDs in November 2014 to 38.3 DIDs in October 2019 (p = 0.002). Selective serotonin reuptake inhibitors were the most sold category of drugs (+5.7 million DDDs) in the period. ‘Other’ antidepressants presented the largest growth rate (104.7%). Individually, the most sold active substance was escitalopram (+1.8 million DDDs), and vortioxetine had the largest growth rate (336.2%). Tricyclic sales remained unchanged, and monoamine oxidase inhibitors had low and even decreasing sales (−9.5%).Discussion: The total sales of antidepressants increased in Brazil from November 2014 to October 2019. The higher sale volumes of selective serotonin reuptake inhibitors and higher growth rate of ‘other’ antidepressants, with low sale volume of tricyclics and a decrease of monoamine oxidase inhibitors, suggest the replacement of older drugs by newer ones following a global trend. Therapeutic advances and commercial promotion efforts on new products might explain these findings. Depression is an important cause of morbidity and disability worldwide . In BrazA growth in antidepressant use is consistent across different settings worldwide . Trends Aggregated information on drug sales may help researchers to understand pharmaceutical market trends and allow comparing different regions into a country or different countries. The World Health Organization (WHO) recommends the Anatomic Therapeutic Chemical (ATC) and the Defined Daily Dose (DDD) methodology, developed by the Norwegian Institute of Public Health, as the gold standard for drug utilization monitoring and research . The DDDMedicalization of life with psychotropic drugs is a recurrent and complex circumstance due to its assignable central role in transforming human conflict and psychological suffering in solvable problems . Drug utDespite the high burden of depressive disorders and feasibility of performing drug utilization studies based on administrative or health services databases, for example, Brazil lacks assessments of antidepressant sales. Pooled data from the pharmaceutical market, including sale registries from drug companies, wholesalers, and retailers, could be useful for estimating the total quantities and trends of antidepressant use in this population. Data from companies specialized in monitoring of the pharmaceutical market, as IQVIA, former IMS Health, may be a source to investigate tendencies of drug sales, in this sense .The aim of this study is to describe the trends of antidepressant sales in Brazil in a five-year period and assess main fluctuations in sales of drugs and pharmacological categories.It is an exploratory ecological study based on the analysis of data of antidepressant sales in Brazil from November 2014 to October 2019.https://consultas.anvisa.gov.br/#/medicamentos/; and Health Environment Legal Prevention & Safety—by Optionline, paid subscription on: https://i-helps.optionline.com/.The data of antidepressant sales in Brazil were provided by IQVIA™, a company that collects daily information on the local pharmaceutical market. The information is based on private pharmacies, hospitals, governments, their wholesaler providers, and direct sales from drug companies. The different sources of sales are complementary and used for cross-checking to avoid double counting. The initial dataset provided data on product name, drug company name, active substance name, European Pharmaceutical Market Research Association (EphMRA) classification code, pharmaceutical form, dosage per package, and quantity of packages sold. The data provided by IQVIA™ contained all N6-coded drugs , which include N6A—antidepressants and mood stabilizers; N6B—psychostimulants; N6C—psycholeptic-psychoanaleptic combinations; N6D—nootropics; N6E—neurotonics; and other miscellaneous products . Only drhttps://www.whocc.no/atc_ddd_index/.We selected all antidepressants in monotherapy presentation. Products with expired market license before the analyzed period were not excluded if there were registries of sales at least in the first studied period. Combination products that included antidepressants in their preparation and mood stabilizers and herbal drugs were excluded. Antidepressants were identified by international nonproprietary name and were recoded according to the ATC classification system. The ATC codes and DDD value data were obtained in March 2020 from the WHO Collaborating Centre for Drug Statistics Methodology website: The primary outcome was the DDD per 1,000 inhabitants per day (DID)  and the To calculate the DID values, we converted the yearly sold package unities of each antidepressant drug to DDD unities. First, the number of DDDs per package was calculated by dividing the total dosage of the active substance per package (in milligrams) by the corresponding DDD value for the substance; then, the result was multiplied by the number of packages of that product sold in each 12-month period of this study . The quantities of products, in DDDs per 12-month period, were pooled by the active substance and by the pharmacological category. To calculate the DID, we considered the Brazilian adult population (≥18 years old) as estimated by the Brazilian Institute of Geography and Statistics for each corresponding year, considering 2015 population for the 2014–5 period and so on .The growth rate was assumed as the percent of change on the quantity of antidepressants sold per substance and per pharmacological category, calculated each 12-month period against the previous 12-month period and the fifth period (2018–9) against the first period (2014–5).The drugs were analyzed both individually and into their respective ATC pharmacological categories: tricyclic antidepressants, named as “non-selective monoamine reuptake inhibitors” at the ATC classification (N06AA); selective serotonin reuptake inhibitors (N06AB); monoamine oxidase inhibitors (N06AF and N06AG); and ‘other’ antidepressants (N06AX), including serotonin–norepinephrine reuptake inhibitors.We described the frequency of sales in DDD and DID and the growth rate in each period. The Chi2 trend value was calculated for assessing whether there was a significant change of DID data between 2014–5 and 2018–9 periods by the active substance, by the pharmacological category, and by the total number of antidepressants sold. Excel Microsoft Office 365 was used for plotting and analyzing the data and for creating tables and figures.Hypericum perforatum), one was a combination (amitriptyline + carbamazepine), and one product based on milnacipran substance was not marketed anymore; the production of products based on moclobemide and reboxetine was discontinued before the analyzed period, but there were sales registered until the third studied period (2016–7). The final dataset for analysis contained 23 active substances in 780 products   . Selecti= 0.002) , and “ot= 0.002) .Total sales of antidepressants increased and sales of tricyclics remained almost unchanged, while those of selective serotonin reuptake inhibitors and ‘other’ antidepressants increased in the period. Monoamine oxidase inhibitors had DID <0.1 in the period .Individually, the main sold antidepressants in terms of the total volume during the five-year period were escitalopram (+1.8 million DDDs), sertraline (+1.35 million DDDs), and fluoxetine (+1.09 million DDDs) . The antThe total sales of antidepressants had increased above two-thirds from November 2014 to October 2019 in Brazil. Selective serotonin reuptake inhibitors had higher sale volume, highlighted by escitalopram, and the ‘other’ antidepressant category had a higher growth rate in the period, mainly due to increase of new marketed drugs such as desvenlafaxine (three-fold increase) and duloxetine (two-fold increase). On the other hand, monoamine oxidase inhibitors experienced sale reduction, especially moclobemide.It is important to consider some limitations of our results. The period of analysis of this study potentially was not enough to analyze associations among the sale trends and possible triggers factors  . The natAntidepressant sales increased in almost 70% in our analysis. Increase in the use of antidepressants from 2014 to 2019 was also observed in other countries, members of the Organization for Economic Co-operation and Development, but at a lower rate: Canada (39%), Chile (20%), Estonia (49%), Iceland (27%), Israel (28%), Latvia (60%), Portugal (35%), and Sweden (17%), among others . The useSelective serotonin reuptake inhibitor sales increased in the period and represented about two-thirds of total antidepressant sales in Brazil. Escitalopram, licensed in 2002 in Brazil, doubled its sales from 2014 to 2019. A similar pattern was observed in Uruguay from 2010 to 2014, with escitalopram as the most used antidepressant . SelectiEscitalopram was the most sold active substance in total volume in Brazil during the assessed period, followed by sertraline and fluoxetine. Most of the antidepressants had their respective generic introduced in Brazil before 2014, but desvenlafaxine and agomelatine had their generics introduced during the studied period, in 2016 and 2018, respectively. Moclobemide, tianeptine, maprotiline, reboxetine, milnacipran, and vortioxetine did not have a generic introduced in the country until October 2019. Sertraline and fluoxetine were also among the most frequently prescribed antidepressants from 1996 to 2015, in the United States, while citalopram and escitalopram were prescribed by an expressive large amount soon after their entry into the market, in 2000 and 2005, respectively . The risIn 2005, the U.S. Food and Drug Administration published a warning for duloxetine due to liver side effects, but it seems not to have caused a prescription decrease in that country . In 2010An apparent gradual replacement of the traditional antidepressants (tricyclics and monoamine oxidase inhibitors) by newer ones (selective serotonin reuptake inhibitors and ‘other’ antidepressants) can be outlined in Brazil. A cohort of employees from Brazilian universities  observedTo the best of our knowledge, this study is a pioneer effort in assessing antidepressant sales in recent years in Brazil. Previous Brazilian assessments of psychotropic drugs relied on data from the Unified Health System , available for the SUS’s specialized component of pharmaceutical care, whose components are newer or more expensive drugs and thus do not represent the therapeutic arsenal for the condition . Our assThe total sales of antidepressants increased from November 2014 to October 2019 in Brazil. The higher sales of selective serotonin reuptake inhibitors and higher growth rate of ‘other’ antidepressants, with low sales of tricyclics and a decrease of monoamine oxidase inhibitors, suggest the replacement of older drugs by newer ones, following a worldwide trend. Further studies may clarify the reasons for the sale trends of these drugs in Brazil.


In all living organisms, extracellular signals are translated into specific responses through signal transduction processes. Research with fungal experimental systems has contributed substantially to uncovering the components participating in these processes, such as signaling molecules, receptors, kinases, phosphatases and second messengers. Different signal transduction pathways interact at various cellular levels and form networks that enable the integration of multiple signals for coordinated cellular responses. Such responses include changes in gene transcription, RNA translation and modification, and protein post-translational and conformational states. Most importantly, these processes are dependent on their cellular locations. These molecular events trigger a variety of fungal cellular reactions and developmental processes, including DNA damage response, cell cycle arrest and progression, pathogenic and symbiotic interactions, asexual and sexual propagation, primary and secondary metabolism, stress adaptation, autophagy, and apoptosis.This special issue includes different research papers and reviews that study the role of signaling molecules, G proteins and receptors, signaling pathways, and multi-subunit complexes allowing the interaction between different signaling pathways, and thus connect upstream regulators and downstream targets. These contributions will promote our understanding of the molecular mechanisms controlling both development and cellular processes in diverse fungal systems .Ganoderma lucidum. They suggest that rebuilding redox homeostasis increases cysteine and proline levels, thus reducing cadmium accumulation. Inositol phosphate signaling is the subject of an article by Murry and coworkers [Schizophyllum commune, they found that altered inositol signaling is involved in tolerance towards metals, such as cesium and zinc, and increased tolerance towards cadmium, which is associated with induced expression of kinases and repression of phosphatases within the inositol cycle. These authors provide a model of how inositol signaling is governed by Ras, G-protein-coupled receptors, and cAMP, and elucidate their different roles in development.Although there is unambiguous evidence that signaling molecules play an essential role in a broad variety of fungal cellular pathways, less is known about the identity of those regulating fungal development. Eight articles contribute to our current knowledge about the molecular players that participate in initiating cellular differentiation. Abdalmegeed et al.  provide oworkers . StudyinMagnaporthe oryzae showed that the molecular mechanisms of fludioxonil action and the perception of osmotic stress are different. In their study, Herrero-de-Dios and co-workers [Candida albicans. In addition to this, a Hog1 mutant strain showed that Hog1 is involved in lipid homeostasis. The authors propose that lipid metabolism contributes to the sensitivity to osmotic stress.Four articles investigated the effect of osmotic stress on sensing. The relationship between external osmotic stress sensing and the fungicidal activity of phenylpyrrole fludioxonil was the subject of a study by Bersching and Jacob . Essenti-workers  showed tMagnaporthe oryzae and addressed the question of how a single gene for a phosphotransfer protein may generate several isoforms to contribute to a multistep phospho-relay system. Their study indicates that alternative splicing is a promising mechanism to generate signal diversity in multistep phospho-relay systems. The HOG pathway is also the subject of a study by Plemenitaš [Hortaea werneckii and the obligate halophilic fungus Wallemia ichthyophaga with the salt-sensitive yeast S. cerevisiae. The author found important structural differences in the HOG pathway components. In conclusion, novel mechanisms of adaptation were driven by changes in salt concentration. In filamentous fungi, the high-osmolarity glycerol (HOG) signaling pathway is present as a multistep phospho-relay system. Bühring et al.  investigemenitaš . Changes2O2, are involved in the signaling of all living organisms and are implicated in a variety of biological processes [2O2 in the ascomycete Aspergillus nidulans. They found that H2O2 affects the phosphorylation of critical regulatory nodes of phosphoinositide, MAPK and TOR signaling, as well as the phosphorylation of multiple proteins involved in diverse developmental processes.Reactive oxygen species (ROS), such as Hrocesses . Carrascrocesses  used a gUstilago maydis and its plant host maize. They investigated extracellular vesicle-associated mRNAs secreted by the maize smut pathogen. Using isolated vesicles, they show by RNA-seq analysis that several mRNAs are enriched and upregulated during infection, and may thus participate in the interaction between the pathogen and host plant.Signaling molecules play a central role in the interspecies or even interkingdom communication of all living organisms. Kwon et al.  are inteSchizosaccharomyces japonicus. Interestingly, strains isolated from diverse host Drosophila species showed a high frequency of sexual sporulation, even under nitrogen-abundant conditions. The authors suggest that their ecological and evolutionary study provides a basis for understanding the sporulation mechanism.Two articles provide quite different experimental systems to understand signal transduction pathways. Groth, Schunke and coworkers  used a fAspergillus fumigatus, lacking a gene for a Gβ-like protein. They showed by organ-specific transcriptional analysis that the Gβ-like protein is essential for completing sexual development. Further roles are associated with phagocytosis by alveolar macrophages, biosynthesis of the cell wall, and oxidative stress responses. A similar experimental approach was used by Tang and coworkers [Botrytis cinerea, which causes gray mold disease in crops. A deletion strain lacking the G protein β subunit gene shows significantly reduced transcription of genes involved in cAMP signaling. From their results the authors conclude that the Gβ protein controls development and virulence through both the cAMP and MAPK signaling pathways.Classical paradigms show that G-protein-coupled receptors (GPCRs) transduce signaling through G proteins. For example, they act as molecular switches in the transduction of intracellular signaling . Since toworkers , on the Aspergillus nidulans. Gene knock-out experiments and fluorescence microscopy data provide evidence for a Golgi-localized TRP-like protein that performs an important function in conidiation, cell wall integration and cellular calcium equilibrium.Transient receptor potential (TRP) proteins are thought to constitute transmembrane ion channels with highly diverse permeation and gating properties. Wang et al.  characteSchizophyllum commune, but the function of other pheromone receptor-like genes has not yet been described. Wirth et al. [S. cerevisiae Ste3a-factor receptor. The data from transcriptome expression analysis and overexpression experiments provide evidence that these genes play a functional role in vegetative growth, possibly by determining growth direction.Pheromone receptor genes are involved in sexual mating of the tetrapolar basidiomycete h et al.  characteAspergillus nidulans. Analyses of the expansion of different families of TFs indicate that the duplication of TFs has an impact at the species level, and the author concludes that the expansion of Zn2Cys6 TFs is mainly due to dispersed duplication events.Signaling pathways activate or silence DNA sequence-specific regulators, such as transcription factors (TFs), which have been intensively studied in members of the Saccharomycotina. In this context, Extebeste  providesThe initial characterization of diverse signaling pathways considered them as linear pathways that do not interact with each other. However, depending on certain physiological conditions, these pathways may enable crosstalk between each other, probably to increase the range of specific cellular responses without increasing the number of components constituting a pathway. The following articles investigated the interplay between two signaling cascades in diverse fungal systems by comparing the phenotypes of signaling-component mutants.Verticillium dahlia. They constructed mutants related to the pheromone response MAPK pathway, or the endoplasmic reticulum-associated unfolded protein response (UPR), both affecting fungal growth, resting structure development, and virulence of the pathogen. The similar phenotypes of signaling-component mutants implied that an interplay exists between UPR and pheromone response MAPK signaling. Similarly, the interplay between two signaling pathways was studied by Moreno-Ruiz et al. [Trichoderma atroviride, a mycoparasitic fungus. The comparative analysis of mutants lacking genes for components of the above-mentioned signaling pathways showed that they are essential for establishing fully functional mycoparasitism.Starke et al.  investigz et al.  They invThe integration of components of several pathways is facilitated by multi-subunit complexes that allow crosstalk between upstream and downstream effectors of the complex . A well-Podospora anserina. For example, the stability of mitochondrial DNA affects lifespan, as does the mitochondrial level of reactive oxygen species (ROS), which has to be balanced since ROS, at low abundance, are essential signaling molecules, but at higher levels cause damage to all kinds of biomolecules. However, when the abundance of ROS passes critical limits, different pathways are induced to cope with the resulting impairments. One of these processes is the degradation of damaged, or excess mitochondria via mitophagy.Mitophagy, the selective vacuolar/lysosomal degradation of mitochondria, is also involved in the aging process, basically occurring in all living beings. The molecular networks that affect this biological process are associated with mitochondria and pathways involved in the biogenesis and quality control of these organelles . The revImproving our understanding of signal transduction networks will require scientific work by many investigators, and I hope that this special issue contributes to this effort. I very much appreciate receiving the articles and reviews from each of the authors, making this special issue possible. Furthermore, I am grateful to the “Journal of Fungi” to have this opportunity to gather articles from the fungal community presenting remarkable work that advances our understanding of “Signal Transductions in Fungi”.