Castration Research Articles

Efficacy and Mechanisms of Oleuropein in Postmenopausal Osteoporosis.

Postmenopausal osteoporosis (PMOP) has a supernal morbidity rate in elderly females. To appraise the effects of oleuropein on bone densitometry, bone metabolic index, oxidative stress, and inflammatory index in PMOP. In addition, the mechanism of olive bittersweet preventing bone loss was explored. We grouped 80 salubrious female Sprague-Dawley rats into four teams: (1) sham operation team (sham, N = 20), (2) ovariectomy (OVX, N = 20), (3) castrated mice fed with oleuropein (OVX+ole, N = 20), and (4) castrated mice fed with estrogen (OVX+E2, N = 20). The ovariectomized SD rats were continuously raised with 200 μg/kg/dose of oleuropein. Bone mineral density and bone metabolism indexes were recorded. In order to assess the effectiveness of oleuropein on osteopenia, an enzyme-linked immunosorbent assay (ELISA) was devoted to examining the bone marrow indexes. The bone metabolism standards of PMOP rats were appraised by assessing serum levels of calcium, alkaline phosphatase (ALP), phosphorus, malondialdehyde (MDA), and nitrate content by experimental detection methods and levels of osteoclastogenesis inhibitory factor (OPG) and receptor activator for nuclear factor-κB ligand (RANKL) by ELISA. The OPG-RANK-RANKL signal passage was examined by Western blot (WB). We measured bone mineral density using dual-energy X-rays. Our animal experimental results indicated that oleuropein could significantly improve the bone mineral density of ovariectomized SD rats. In the meantime, it could reduce ending interleukin-6 (IL-6), malondialdehyde (MDA), nitrate, alkaline phosphatase (ALP), and phosphorus (P) serum concentration and would not affect Ca2+ concentration. In cell experiments, oleuropein also can promote the proliferation of osteoblasts. Furthermore, it can promote the expression of OPG protein and mRNA. In reverse, it inhibits the expression of RANKL protein and mRNA. Oleuropein can not only improve the inflammatory and oxidative indexes of castrated rats but also prevent osteoporosis. Oleuropein avoids bone resorption by regulating OPG/RANKL expression.

Measurement challenges and causes of incomplete results reporting of biomedical animal studies: Results from an interview study.

BackgroundExisting evidence indicates that a significant amount of biomedical research involving animals remains unpublished. At the same time, we lack standards for measuring the extent of results reporting in animal research. Publication rates may vary significantly depending on the level of measurement such as an entire animal study, individual experiments within a study, or the number of animals used.MethodsDrawing on semi-structured interviews with 18 experts and qualitative content analysis, we investigated challenges and opportunities for the measurement of incomplete reporting of biomedical animal research with specific reference to the German situation. We further investigate causes of incomplete reporting.ResultsThe in-depth expert interviews revealed several reasons for why incomplete reporting in animal research is difficult to measure at all levels under the current circumstances. While precise quantification based on regulatory approval documentation is feasible at the level of entire studies, measuring incomplete reporting at the more individual experiment and animal levels presents formidable challenges. Expert-interviews further identified six drivers of incomplete reporting of results in animal research. Four of these are well documented in other fields of research: a lack of incentives to report non-positive results, pressures to ‘deliver’ positive results, perceptions that some data do not add value, and commercial pressures. The fifth driver, reputational concerns, appears to be far more salient in animal research than in human clinical trials. The final driver, socio-political pressures, may be unique to the field.DiscussionStakeholders in animal research should collaborate to develop a clear conceptualisation of complete reporting in animal research, facilitate valid measurements of the phenomenon, and develop incentives and rewards to overcome the causes for incomplete reporting.

Unilateral Tb Epididymo-Orchitis with Scrotal Fistula- A Case Report

Tuberculosis is one of the most common worldwide diseases that remain almost unaltered in prevalence for the past few years. Genitourinary tract tuberculosis is common, but when associated with scrotal fistula it is a rare presentation. A 50 years old male came to the hospital with the complaints of pricking pain and swelling in the left side of scrotum for the past 2 months. He had history of smoking for past 40 years, on examination swelling of size approximately 6x5 cm present in left side of scrotum, globular in shape; penis pushed to right side was present, lower part of the swelling was firm to hard in consistency, left cord thickness was present and fixity to skin was also present. There was a small fistulous opening present in the left side of scrotum with no active discharge. Bilateral inguinal lymph nodes were palpable. This is a rare case presentation of tuberculous epididymo-orchitis with scrotal fistula leading to testis removal. Initially emperical treatment was started for TB epididymoorchitis which turned unresponsive, so he was then proceeded for surgery to avoid further disease progression. This case was managed radiologically, clinically and pathologically. This case report shows how the case was managed and followed up.Since symptoms of TB epididymo-orchitis are milder when compared to the symptoms of pyogenic epididymo-orchitis, they appear to physicians in later advanced stages. Tuberculosis should be considered in Patients with mild symptoms of epididymo-orchitis, mainly in those with HIV, immunosuppressive conditions and endemic areas.

Preparation and Evaluation of Siderol Amorphous Solid Dispersions: Selection of Suitable Matrix/Carrier.

The present study investigates the preparation of amorphous solid dispersions (ASD) for the ent-kaurane diterpenoid siderol (SDR). Initially, evaluation of the pure drug (isolated from Sideritis scardica) revealed that the API is a non-stable glass former, and hence the selection of a suitable ASD's matrix/carrier needs special attention. For this reason, four commonly used polymers and copolymers, namely poly(vinylpyrrolidone), copovidone, hydroxypropyl cellulose, and Soluplus® (SOL), were screenedviafilm casting and crystal growth rate measurements. Amongst them, SOL showed the highest SDR's crystal growth rate reduction, and, since it was also miscible with the drug, it was selected for further testing. In this direction, SDR-SOL ASDs were successfully preparedviamelt-quench cooling. These formulations showed full API amorphization, while good physical stability (i.e., a stable SDR amorphous dispersions) were obtained after storage for several months. Finally, evaluation of molecular interactions (with the aid of ATR-FTIR spectroscopy) showed strong H-bonds between SOL and SDR, while the use of molecular dynamics (MD) simulations unraveled the nature of these interactions. Therefore, based on the findings of the present work, SOL seems to be an appropriate matrix/carrier for the preparation of SDR ASDs, although further studies are needed in order to explore its full potentials.

Features of formation of the structure of thin slabs cast at different speeds, their influence on the structure and properties of hot-rolled steel

For the first time, a complete comprehensive study of the internal quality of metal products with a carbon content of 0.3 % produced in a combined process of thin-slab casting and rolling was carried out. A feature of the combined process is the absence of a cooling and heating cycle before rolling and the small thickness of the initial slab. The zonal chemical heterogeneity of continuously cast "thin" slabs poured at different speeds has been studied. The research methods were metallographic analysis of the macrostructure and spectral method for determining the content of elements by the thickness of slabs. The results of calculations of the segregation coefficients of basic and impurity chemical elements in slabs poured at different speeds are presented. As a result of the study, it was found that an increase in the casting rate by ~ 1.2 times does not lead to a significant increase in the zonal chemical heterogeneity of the continuously cast metal produced in the process of thin-slab casting. Of particular interest was the measurement of quantitative parameters of the initial dendritic structure of "thin" slabs depending on the casting speed. The segregations formed as a result of dendritic liquation were measured. It is shown that with an increase in the casting rate, the dispersion of the dendritic structure increases and the dendritic segregation decreases by 20 %. In the course of simulation rolling, the regularity in changing the parameters of the dendritic structure during deformation of "thin" slabs into the final microstructure of rolled products is considered. When the slabs are compressed by 65 %, the distance between the primary dendrite arm spacing decreases by an average of 65 %. The comparison of the microstructure and mechanical characteristics of rolled products produced in industrial conditions from slabs poured at different speeds using thin-slab casting technology is carried out. The results of the study indicate that an increase in the casting rate increases the dispersion of the initial dendritic structure and, as a result, favorably affects the formation of the final microstructure. It is shown that hot-rolled products made from a slab with a 1.2-fold increased casting speed have a homogeneous microstructure and demonstrate an increase in the values of impact strength by an average of 30 J/cm2.

Transcriptome-proteome compendium of the Antarctic krill (Euphausia superba): Metabolic potential and repertoire of hydrolytic enzymes.

The Antarctic krill (Euphausia superba Dana) is a keystone species in the Southern Ocean that uses an arsenal of hydrolases for biomacromolecule decomposition to effectively digest its omnivorous diet. The present study builds on a hybrid-assembled transcriptome (13,671 ORFs) combined with comprehensive proteome profiling. The analysis of individual krill compartments allowed detection of significantly more different proteins compared to that of the entire animal (1464 vs. 294 proteins). The nearby krill sampling stations in the Bransfield Strait (Antarctic Peninsula) yielded rather uniform proteome datasets. Proteins related to energy production and lipid degradation were particularly abundant in the abdomen, agreeing with the high energy demand of muscle tissue. A total of 378 different biomacromolecule hydrolysing enzymes were detected, including 250 proteases, 99 CAZymes, 14 nucleases and 15 lipases. The large repertoire in proteases is in accord with the protein-rich diet affiliated with E. superba's omnivorous lifestyle and complex biology. The richness in chitin-degrading enzymes allows not only digestion of zooplankton diet, but also the utilisation of the discharged exoskeleton after moulting.

Modeling of changes in the cerebral cortex under low radiation exposure

The aim of the investigation was to reveal changes in the neurons of the cerebral cortex depending on the dose of γ-irradiation, the dose rate of exposure, the mode of exposure (acute and chronic) and to establish the priority of each of the influencing factors (γ-irradiation, recovery time and their joint impact) in the end result.Material and methods. The experiment, in compliance with the rules of bioethics, was carried out on 300 adult white rats that were exposed to acute and chronic γ-irradiation in total doses from 10 to 100 cGy with a dose rate of 50 to 660 cGy/h. The sensory and motor areas of the cerebral cortex were studied throughout life after exposure to radiation using traditional neuromorphological, histochemical, and statistical methods, followed by mathematical modeling of the obtained evaluation criteria.Results. Despite the peculiarities of the regimes of radiation exposure, all of them did not cause statistically significant changes in functionally different areas of the cerebral cortex. Throughout the entire recovery period, both in control and experimental animals, phase stochastic changes were observed in various neuromorphological indicators of the state of nerve cells, which, as a rule, had a borderline character, and the level of their significance fluctuated in relation to the control within insignificant limits. At the same time, the change in some neuromorphological parameters was more influenced by the dose or dose rate of γ-irradiation, while others were influenced by the recovery time. With the combined effect of the radiation factor and the duration of the recovery period, most of the indicators corresponded to the age control.Conclusion. Statistically significant changes in the state of neurons in both sensory and motor areas of the cerebral cortex under the considered modes of γ-irradiation in total doses up to 100 cGy were not revealed.

Cross Sectional Survey of Canine Idiopathic Epilepsy Management in Primary Care in the United Kingdom.

The aims of this study are to gain insight on how primary care veterinarians in the UK diagnose and treat canine idiopathic epilepsy (IE) and what they perceive as challenges in the management of canine IE. Two hundred and thirty-five primary care veterinarians took part in this survey. The questionnaire asked about the type of practice the respondent worked in, any relevant post-graduate qualifications, how many years' experience they had in practice and the participant's canine IE caseload. Participants were asked how they diagnose canine IE, how they select antiseizure drugs (ASDs) and how they assess outcome. The questionnaire also explored which information sources they have access to for deciding on canine IE treatment, challenges that may be faced when managing these cases and areas in which more support can be provided. 94.5% of participants (n = 222/235) managed <10 canine IE cases in a year and 87.8% (n = 206/235) used phenobarbital as their first line ASD. The reported mean initial phenobarbital dose was 2.1 mg/kg (standard deviation = 0.71) every 12 h. When considering how closely participants aligned with IVETF guidelines on the topics of diagnosis, ASD initiation and outcome assessment, on average participants would score around half of the available points. 53.2% (n = 125/235) of respondents recommended neutering in canine IE and 46.8% (n = 110/235) did not. 53.2% (n = 125/235) did not recommend any additional treatments for canine IE beyond use of ASDs. 23.4% recommended Purina Neurocare diet (n = 55/235), 12.8% recommended environmental modification (n = 30/235), and 6.8% (n = 16/235) recommend medium chain triglyceride supplements. In this study participants found managing client expectations to be most challenging alongside canine IE emergency management. The main limitation of this study is the relatively low response rate and therefore the results may not reflect the entire small animal veterinary profession in the UK. However, the results of this study represent a starting point to inform educational resources and support strategies to improve quality care of canine IE in primary care.

Androgenic properties of “Dong Trung Ha Thao Sapa” hard capsules on experimental animals

“Dong trung ha thao Sapa” (DTHT) hard capsules prepared from Cordyceps militaris powder are intended to treat male hypogonadism. The aim of the current study was to evaluate the androgenic properties of this product on experimental animals. Weanling male rats and castrated peripubertal male rats were used for the study. Weanling male rats were divided into 5 groups: group I (saline-treated); group II (testosterone undecanoate-treated); group III (low dose DTHT capsules); and group IV (high dose DTHT capsules). Castrated peripubertal male rats were also divided into 5 groups: group I without castration (saline-treated); groups II to V were surgically castrated, group II (saline-treated); group III (testosterone undecanoate-treated), group IV (low dose DTHT capsules) and group V (high dose DTHT capsules). The potential effect was evaluated on five androgen-dependent tissue weight (seminal vesicles, ventral prostate, Cowper’s gland, glans penis, levator ani-bulbocavernosus - LABC muscle and the serum testosterone level in castrated rats. Five tissue weights and testes, epididymis weights were examined in weanling rats. A positive androgen agonist result should be a statistically significant increase (p ≤ 0.05) in any two or more of the five targets’ androgen-dependent tissue. The results suggest that capsules show androgenic properties in castrated male rats by increasing weight of 3 tissues (seminal vesicles, ventral prostate, LABC muscle). DTHT hard capsules appear to be an effective therapeutic drug for treating hypogonadism.

Origanum majorana L. Extract Attenuated Benign Prostatic Hyperplasia in Rat Model: Effect on Oxidative Stress, Apoptosis, and Proliferation.

Benign prostatic hyperplasia (BPH) is a widespread androgenic illness influencing elderly men. It is distinguished by prostatic epithelial and stromal muscle cell proliferation. Inflammation, oxidative stress, and apoptosis have all been interrelated to the development of BPH. Marjoram (Origanum majorana L.) is a herb with reported antiproliferative, proapoptotic, and antioxidative properties, which have not yet been studied in relation to BPH. Consequently, in this work, an ethanolic extract of O. majorana was prepared in two doses (250 and 500 mg/kg/day) to be injected into castrated rats after induction of a testosterone-BPH model. Testosterone propionate (TP) was subcutaneously injected (0.5 mg/kg/day) for one week after castration to induce BPH. Forty adult Wistar male rats were randomly allocated into five groups: control, BPH model, high and low O. majorana doses (250, 500 mg/kg/day), and finasteride (FN) (0.8 mg/kg/day) as a positive control. Treatment was continued with drugs/normal saline for 28 days. Rat’s body and prostate were weighed, prostate index (PI) and % of prostate growth inhibition were calculated, serum dihydrotestosterone (DHT), prostatic content of superoxide dismutase (SOD), catalase (CAT), total antioxidant capacity (TAC), and malondialdehyde (MDA), DN damage, histopathological changes, immune expression of proliferating cell nuclear antigen (PCNA), caspase-3, α-SMA, and TGF-β1 were assessed. In addition, molecular quantitative PCR and ELISA analyses were performed to identify the expression of mRNAs and related proteins of both caspase-3 and TGF-β1 in prostate tissue from O. majorana-treated and untreated groups. Rats with BPH had significantly higher prostate weights and PI, higher DHT, DNA damage (8-hydroxyguanine, 8-OH-dG), and MDA levels with prominent PCNA, α-SMA, and TGF-β expression, but lower SOD, CAT, and TAC activity and caspase-3 expression. O. majorana (250 and 500 mg/kg/day)-treated groups revealed a decrease in prostate weights and PI, lower levels of DHT, suppressed oxidative stress, reduced tissue proliferation and fibrosis, and restored antioxidant and proapoptotic activity. Additionally, quantitative PCR and ELISA analysis showed that treatment with O. majorana significantly upregulated the expression of caspase-3 and downregulated the expression of TGF-β in prostate tissues of BPH rats. The data were confirmed by the immunohistological reactivity of these targeted markers in the prostate tissues. These effects were more significant with O. majorana 500 mg/mL/rat. In conclusion, the current study indicates the efficient use of O. majorana in the treatment of testosterone-induced BPH through its antiproliferative, proapoptotic, and antioxidative mechanisms.

053 Totally extraperitoneal approach (TEP) for gonadectomy for differences in sex development (DSD): Report of 2 cases

Introduction: 17-β-hydroxysteroid dehydrogenase type-3 deficiency (17βHSD-3) is a rare cause of 46XY DSD. Gonadectomy is often offered to prevent further virilization and to continue living in a female gender role [ [1] Chuang J. Vallerie A. Breech L. Saal H.M. Alam S. Crawford P. Rutter M.M. Complexities of gender assignment in 17β-hydroxysteroid dehydrogenase type 3 deficiency: is there a role for early orchiectomy?. Int J Pediatr Endocrinol. 2013; 12: 15 Crossref Google Scholar ]. Both laparotomic and laparoscopic approaches to gonadectomy had been described for DSDs [ [2] Dellaportas D. Chatzipapas I. Pandraklakis A. Michala L. Laparoscopic bilateral gonadectomy and inguinal hernia repair with mesh for complete androgen insensitivity syndrome: A case report. J Pediatr Adolesc Gynecol. 2019; 32 (Epub 2019 Feb 27): 443-445 Abstract Full Text Full Text PDF PubMed Scopus (1) Google Scholar ]. Retrieval and removal of testes located in the inguinal canal can be technically challenging and requires collaboration with surgical colleagues. Cases: We present 2 cases of DSD with 17βHSD-3. One of them is 16 years old, while the other one is 20 years old. Both cases presented with amenorrhea and clitoral enlargement, whereas the second case also had deepening of voice. Blood tests for both revealed elevated testosterone and androstenedione. MRI findings were similar showing bilateral gonads located in the inguinal canals. Genetic tests revealed 46 XY karyotype and HSD17B3 gene mutation. Both subjects opted to continue with a female gender role and opted for gonadectomy to prevent further virilization. Gonadectomy was performed with a totally extraperitoneal laparoscopic approach. An infraumbilical incision was made and care was taken to enter the potential space above the rectus sheath. Then a balloon was placed via the port to distend the extraperitoneal space and then the laparoscope was inserted. Secondary ports at the suprapubic region and the left lower quadrant were placed. The gonads were mobilized and gonadectomy performed. Gonadectomy was successfully performed for the first case, but for the second case, there was failure to mobilize the gonad and operation was completed with inguinal incisions. Conclusion: Totally extraperitoneal laparoscopic approach for hernia repair is a well-established technique familiar to surgeons [ [3] McKernan J.B. Laws H.L. Laparoscopic repair of inguinal hernias using a totally extraperitoneal prosthetic approach. Surg Endosc. 1993; 7: 26-28 Crossref PubMed Scopus (264) Google Scholar ]. This was the first time the TEP approach has been described for gonadectomy for DSD. By cooperating with surgeons who has abundant experience in operating in this region, one can allow utilization of techniques which were already commonly used. It raises the importance as gynecologists, we should be mindful of technological advances in other specialties as with other aspects of DSD, collaboration with other specialties may lead to advances in patient management.

FELASA-AALAS Recommendations for Biosecurity in an Aquatic Facility, Including Prevention of Zoonosis, Introduction of New Fish Colonies, and Quarantine.

FELASA and AALAS established a joint working group to advise on good practices for the exchange of fish for research. In a first manuscript, the working group made recommendations for health monitoring and reporting of monitoring results. The focus of this second related manuscript is biosecurity in fish facilities. First, we define the risk of contamination of personnel by zoonotic pathogens from fish or from system water, including human mycobacteriosis. Preventive measures are recommended, such as wearing task-specific personal protective equipment. Then we discuss biosecurity, highlighting the establishment of biosecurity barriers to preserve the health status of a facility. A functional biosecurity program relies on integration of the entire animal facility organization, including the flow of staff and animals, water treatments, and equipment sanitation. Finally, we propose 4 steps for introducing new fish colonies: consideration of international trade and national restrictions; assessing risk according to fish source and developmental stage; establishing quarantine barriers; and the triage, screening, and treatment of newly imported fish. We then provide 3 realistic sample scenarios to illustrate practical biosecurity risk assessments and mitigation measures based on considerations of health status and quarantine conditions.

Testosterone Influence on Microtubule-Associated Proteins and Spine Density in Hippocampus: Implications on Learning and Memory

The thorny protrusions or spines increase the neuronal surface area, facilitate synaptic interconnections among neurons, and play an essential role in the hippocampus. Increasing evidence suggests that testosterone, the gonadal hormone, plays an important role in neurogenesis and synaptic plasticity. The role of testosterone on microtubule-associated proteins on dendritic neurite stability in the hippocampus and its impact on learning disability is not elucidated. Adult male Wistar albino rats were randomly selected for the control, castrated, castrated + testosterone, and control + testosterone groups. Bilateral orchidectomy was done, and the testosterone propionate was administered during the entire trial period, i.e., 14 days. The learning assessments were done using working/reference memory versions of the 8-arm radial maze and hippocampal tissues processed for histological and protein expressions. There were reduced expressions of microtubule-associated protein 2 (MAP2), postsynaptic density protein 95 (PSD95), and androgen receptor (AR) and increased expression of pTau in the castrated group. Conversely, the expression of MAP2, PSD95, and AR was increased, and the pTau expression was reduced in the hippocampus of the castrated rat administrated with testosterone. Androgen-depleted rats showed impaired synaptic plasticity in the hippocampus associated with contracted microtubule dynamics. Along with learning disability, there was an increased number of reference memory errors and working memory errors in castrated rats. Observations suggest that androgen regulates expression of neural tissue-specific MAPs and plays a vital role in hippocampus synaptic plasticity and that a similar mechanism may underlie neurological disorders in aging and hypogonadal men.

Effect of androgen replacement therapy on erectile function through histological recovery in castrated rats

ABSTRACT Introduction Androgens are essential for physical activity in males and normal erectile function. Reportedly, androgen deficiency causes erectile dysfunction through tissue damage in the corpus cavernosum. We previously reported that delayed androgen replacement therapy improved erectile function in rats through histological recovery. Objective To investigate the mechanism of the recovery process by androgen replacement therapy in rats. Methods Twelve-week-old male Wistar ST rats were used in this study. The rats were assigned to the following groups: castrated (Cast), castrated with subcutaneous administration of testosterone undecanoate (25 mg/kg; Cast+T), and sham (Sham). Cast+T rats received subcutaneous doses of testosterone, beginning from 4 weeks after castration, whereas Sham and Cast rats received only the vehicle. Erectile function was assessed by evaluating intracavernosal pressure (ICP) and mean arterial pressure (MAP) after electrical stimulation of the cavernous nerve at 1, 2, 4, and 8 weeks after testosterone administration. mRNA expression of histological factors (Cavin-1, Cavin-2, Cavin-3, α-SMA, VEGF-R, ICAM-1, and TGF-β1) was tested using real-time PCR. Results The ratio of ICP to MAP was 0.26 ± 0.04 in the 4th week for the Cast group, and the erectile function was significantly lower than that of the Sham group (0.61 ± 0.03; p &amp;lt;0.05). On the other hand, in the Cast+T group, it changed over time from 0.41 ± 0.06 in the 1st week, 0.44 ± 0.04 in the 2nd week, 0.48 ± 0.04 in the 4th week, to 0.70 ± 0.03 in the 8th week, and erectile function was significantly improved 8 weeks after the testosterone administration (p &amp;lt;0.05). The Cavin-1 and Cavin-2 mRNA expressions were increased in the Cast+T group compared to the Cast group at all times (P &amp;lt;0.01). α-SMA, VEGF-R, ICAM-1, and TGF-β1 mRNA expressions were also increased in the Cast+T group, as compared to the Cast group (P &amp;lt;0.01). Conclusions In this study, we identified that androgen replacement therapy in rats would improve ED. This may be because the administration of testosterone improved histological recovery. It has been reported that these caveolae-related factors control the invagination structure of the cell membrane, and that caveolae in endothelial cells are also involved in the production of nitric oxide (NO), which might contribute to the therapeutic effect of androgen replacement therapy. Increase in the expression of these caveolae-related factors may regenerate smooth muscle and corpus cavernosum structures. Disclosure Work supported by industry: no.

Castration mediated schwann cell dedifferentiation leads to slower nerve conduction, decreased neuritogenesis, and nitrergic neuron loss

ABSTRACT Introduction/Objectives Low levels of androgens due to androgen deprivation therapy for prostate cancer treatment or due to aging lead to erectile dysfunction (ED). Castration, a preclinical model of low androgens, induces ED due to damage to pelvic nerves and a loss of erectogenic nitrergic nerves. Schwann cells (SC) release neurotrophic factors to promote pelvic nerve growth and survival. It is unknown how testosterone deficiency impacts SC function. This study determines how testosterone deficiency influences SC phenotype and trophic support in neuronal regeneration. We hypothesize testosterone deficiency will decrease myelinated pelvic SC populations, delay penile nerve conduction and reduce pelvic nitrergic neurons. Methods Male Sprague Dawley rats (12 wks) were divided into control (CON) and surgical castration (CAST) groups. All experiments were performed six weeks following castration. Dorsal penile nerves (DPN; 15-16 mm) were excised, placed in a sylgard dish containing oxygenated buffer and conduction velocity of myelinated fast fibers was assessed (n=8 per group). The major pelvic ganglia (MPG) were collected and dissociated into SC or neurons. Dissociated neurons were grown on laminin coated coverslips. To determine whether SCs could provide trophic support, we co-cultured neurons on top of confluent SCs. We assessed the following co-culture groups: CON SC + CON MPG, CON SC + CAST MPG, CAST SC + CON MPG (n=4 per group, cultured in triplicate). After 72 hours, all cultures were fixed and stained with beta tubulin class III, neuronal nitric oxide synthase (nNOS), and tyrosine hydroxylase (TH) to assess neurite length and branching, nitrergic and sympathetic neuron populations, respectively. Dissociated SC were characterized via qPCR for markers of all SC (Sox10), myelinated SC (Krox20), and dedifferentiated SC (glial fibrillary acidic protein; GFAP). Results CAST slows DPN myelinated fast fiber conduction velocity. In dissociated cultured MPG neurons, castration reduces both neurite length and number of branches. Interestingly, CAST MPG neurons co-cultured with CON SC restore neurite length and branching to same level as CON MPG neurons co-cultured with CON SC; however, CAST SC do not impact CON MPG neurite length. CAST MPG neurons co-cultured with CON SC and CON MPG neurons co-cultured with CAST SC both have less nitrergic neurons than CON SC + CON MPG neurons. CAST does not impact the number of TH neurons in single or co-cultured conditions. CAST reduces gene expression of Sox10, myelinated SC, and dedifferentiated SC. Conclusions CAST markedly impairs penile nerve conduction, reduces neurite outgrowth and the proportion of pro-erectile nitrergic neurons. Healthy SC are able to rescue neuritogenesis but not the survival of nitrergic neurons. Additionally, healthy MPG neurons plated on CAST SC have a reduced number of nitrergic neurons. We believe CAST SC are undergoing a phenotypic switch from a myelinated neurotrophic form into dedifferentiated SC promoting neuritogenesis and loss of nitrergic neurons. Preserving the myelinated SC phenotype may prevent impaired nerve damage and subsequent ED in patients with testosterone deficiency. Disclosure Work supported by industry: no.

Aortic Relaxation in Castrated Male Rats and Responses to Estrogen Treatment

Previous studies suggest that risk factors for cardiovascular diseases (CVD) have increased in the transgender population after they undergo cross‐sex hormone therapy (CSHT). Several studies, including ours in animal models as well as in humans, have shown that estrogen exerts direct beneficial effects on the vessel wall in healthy females. A growing body of evidence also suggests that androgen exhibits protective actions on the cardiovascular system. However, the effects of CSHT in cardiovascular physiology in males or females are less understood.The current study was, therefore, undertaken to evaluate the effects of estrogen treatment on aortic reactivity in castrated (CAS) male rats. Specifically, we investigated the effects of in vivo treatment with 17β‐estradiol (E2) on the aortic responses of CAS male as well as ovariectomized (OVX) female rats.Age‐matched CAS male and OVX female rats were implanted subcutaneously with either 17β‐estradiol (E2,1.5 mg) or a placebo‐containing pellet for 30 days. Endothelium‐dependent vasorelaxation (EDV) in response to acetylcholine (ACh, 10‐8 to 10‐5 M) in aorta precontracted with phenylephrine (PE, 2 µM) was measured before and after pretreatment with indomethacin (10 µM, cyclooxygenase inhibitor), followed by N‐nitro‐ L‐arginine methyl ester [L‐NAME, 200 µM, a nonselective nitric oxide synthase (NOS) inhibitor]. Endothelium‐independent vasorelaxation was assessed by measuring sodium nitroprusside (SNP)‐induced relaxation response in endothelium‐denuded rings in addition to the contractile response curves to PE before and after pretreatment with L‐NAME in aortic rings.EDV to ACh was significantly impaired in aortic rings from E2‐treated CAS rats compared to their placebo‐treated counterparts and E2‐treated OVX rats. The impairment of EDV to ACh in E2‐treated CAS rats was accompanied by a significant decrease in nitric oxide (NO) contribution to ACh‐induced relaxation in this group. The smooth muscle sensitivity to NO as measured by SNP relaxation response curve was not altered, whereas the responsiveness to PE was slightly enhanced in the aortas of E2‐treated CAS group compared to those in other experimental groups. Endothelium‐derived NO release during smooth muscle contraction, as assessed by the potentiation of the response to PE after NOS inhibition, was not different in the aortas of experimental groups.Overall, our data demonstrate that aortic function was altered in E2‐treated CAS male rats. Here, we provide the first evidence of impairment in aortic relaxation in E2‐treated CAS male rats, possibly via a significant decrease in the relative contribution of NO to the regulation of vascular relaxation response. Furthermore, the comparison of current data in the CAS male rats with our recently published data on the aortic reactivity in the intact males appears to rule out any effect of testosterone deprivation on the stimulated release of NO in male rat aortas. Additional studies will be needed to document the direction and magnitude of CSHT interaction with vasculatures.

HFC electric-welded pipes made of structural steel with resistance to hydrogen cracking

The results of studies carried out in laboratory and industrial conditions, the influence of an integrated approach consisting in the modification of liquid steel alkaline earth metals and rare earth metal, as well as double heat treatment (hardening from the austenitic region at 920 &ordm;C and subsequent high tempering at 650 &ordm;C) of pipes welded with high frequency currents from structural steel with a content of 0.18&ndash;0.22 % carbon on the resistance against cracking in H2S-containing media according to the NACE TM 0284 standard are presented. It is shown thatthe adjustment of the integrated approach in terms of limiting the casting rate made it possible to completely eliminate the formation of cracks of hydrogen origin in the metal (CLR = 0 %, CTR = 0 %). A technology for the production of rolled products and pipes welded with high frequency currents with requirements for cold resistance and corrosion resistance in environments containing hydrogen sulfide has been developed. The cost of pipe products made of structural steel with 0.18&ndash;0.22 % C, produced according to the developed technology, is less than 2000 rubles/ton compared to similar pipe products made of steel grades 05HGB, 13HFA, 09GSF.

HYBRiD: hydrogel-reinforced DISCO for clearing mammalian bodies.

The recent development of solvent- and polymer-based brain-clearing techniques has advanced our ability to visualize the mammalian nervous system in three dimensions. However, it remains challenging to image the mammalian body en bloc. Here we developed HYBRiD (hydrogel-based reinforcement of three-dimensional imaging solvent-cleared organs (DISCO)), by recombining components of organic- and polymer-based clearing pipelines. We achieved high transparency and protein retention, as well as compatibility with direct fluorescent imaging and immunostaining in cleared mammalian bodies. Using parvalbumin- and somatostatin-Cre models, we demonstrated the utility of HYBRiD for whole-body imaging of genetically encoded fluorescent reporters without antibody enhancement of signals in newborn and juvenile mice. Using K18-hACE2 transgenic mice, HYBRiD enabled perfusion-free clearing and visualization of SARS-CoV-2 infection in a whole mouse chest, revealing macroscopic and microscopic features of viral pathology in the same sample. HYBRiD offers a simple and universal solution to visualize large heterogeneous body parts or entire animals for basic and translational research.

Connexinplexity: the spatial and temporal expression of connexin genes during vertebrate organogenesis.

Animal development requires coordinated communication between cells. The Connexin family of proteins is a major contributor to intercellular communication in vertebrates by forming gap junction channels that facilitate the movement of ions, small molecules, and metabolites between cells. Additionally, individual hemichannels can provide a conduit to the extracellular space for paracrine and autocrine signaling. Connexin-mediated communication is widely used in epithelial, neural, and vascular development and homeostasis, and most tissues likely use this form of communication. In fact, Connexin disruptions are of major clinical significance contributing to disorders developing from all major germ layers. Despite the fact that Connexins serve as an essential mode of cellular communication, the temporal and cell-type-specific expression patterns of connexin genes remain unknown in vertebrates. A major challenge is the large and complex connexin gene family. To overcome this barrier, we determined the expression of all connexins in zebrafish using single-cell RNA-sequencing of entire animals across several stages of organogenesis. Our analysis of expression patterns has revealed that few connexins are broadly expressed, but rather, most are expressed in tissue- or cell-type-specific patterns. Additionally, most tissues possess a unique combinatorial signature of connexin expression with dynamic temporal changes across the organism, tissue, and cell. Our analysis has identified new patterns for well-known connexins and assigned spatial and temporal expression to genes with no-existing information. We provide a field guide relating zebrafish and human connexin genes as a critical step toward understanding how Connexins contribute to cellular communication and development throughout vertebrate organogenesis.

The enteric nervous system of the C. elegans pharynx is specified by the Sine oculis-like homeobox gene ceh-34.

Overarching themes in the terminal differentiation of the enteric nervous system, an autonomously acting unit of animal nervous systems, have so far eluded discovery. We describe here the overall regulatory logic of enteric nervous system differentiation of the nematode Caenorhabditis elegans that resides within the foregut (pharynx) of the worm. A C. elegans homolog of the Drosophila Sine oculis homeobox gene, ceh-34, is expressed in all 14 classes of interconnected pharyngeal neurons from their birth throughout their life time, but in no other neuron type of the entire animal. Constitutive and temporally controlled ceh-34 removal shows that ceh-34 is required to initiate and maintain the neuron type-specific terminal differentiation program of all pharyngeal neuron classes, including their circuit assembly. Through additional genetic loss of function analysis, we show that within each pharyngeal neuron class, ceh-34 cooperates with different homeodomain transcription factors to individuate distinct pharyngeal neuron classes. Our analysis underscores the critical role of homeobox genes in neuronal identity specification and links them to the control of neuronal circuit assembly of the enteric nervous system. Together with the pharyngeal nervous system simplicity as well as its specification by a Sine oculis homolog, our findings invite speculations about the early evolution of nervous systems.