Impacts of DES on the expressions of related genes in the gubernaculums testis of newborn mice / 中华男科学杂志

Objective@#To investigate the influence of diethylstilbestrol (DES) on the mRNA expressions of the androgen receptor (AR), estrogen receptor α (ERα), proliferating cell nuclear antigen (PCNA), and actin alpha 1 (ACTα1) in the gubernaculums testis of newborn mice and explore their action mechanisms.@*METHODS@#A total of 140 male Kunming mice were randomly divided into a blank control, a dimethyl sulfoxide (DMSO) control, and 5 experimental groups to be treated subcutaneously with normal saline, DMSO, and DES at 0.02, 0.1, 0.5, 10 and 50 μg per kg of the body weight per day, respectively, at gestation days 9－17. On the first day after birth, the animals were sacrificed and the gubernaculums testis collected for detection of the mRNA expressions of AR, ERα, PCNA and ACTα1 by RT-PCR.@*RESULTS@#Compared with the DMSO control, the experimental groups, particularly the DES 10 and 50 μg groups, showed significant increases in the mRNA expression of ERα (RE2 = 0.825, P <0.05), but remarkable decreases in those of AR, PCNA and ACTα1 (RA2 = 0.713, RP2 = 0.946, RT2 = 0.960, P <0.01), all in a dose-dependent manner.@*CONCLUSIONS@#The AR, ERα, PCNA, and ACTα1 mRNA are expressed in the gubernaculum testis of normal newborn mice, and their expression levels may be influenced by intervention with different concentrations of DES during the gestation. Exogenous estrogens may affect the proliferation and contraction of gubernaculum testis cells and consequently the normal development of the testis or even the whole male reproductive system by influencing the metabolism of ER and/or AR.

Roles of G protein-coupled estrogen receptor in the male reproductive system / 中华男科学杂志

The G protein-coupled estrogen receptor (GPER), also known as G protein-coupled receptor 30 (GPR30), was identified in the recent years as a functional membrane receptor different from the classical nuclear estrogen receptors. This receptor is widely expressed in the cortex, cerebellum, hippocampus, heart, lung, liver, skeletal muscle, and the urogenital system. It is responsible for the mediation of nongenomic effects associated with estrogen and its derivatives, participating in the physiological activities of the body. The present study reviews the molecular structure, subcellular localization, signaling pathways, distribution, and function of GPER in the male reproductive system.

Diethylstilbestrol affects LGR8 expression in mouse gubernaculum testis / 中华男科学杂志

<p><b>OBJECTIVE</b>To investigate the impact of prenatal exposure to diethylstilbestrol (DES) on the specific receptor LGR8 of insulin-like factor 3 (INSL3) in the mouse gubernaculum testis, and that of exoestrogens on descensus testis in mice.</p><p><b>METHODS</b>A total of 120 pregnant KM mice aged 8 to 10 weeks were assigned to a normal, a blank control and 4 DES groups of equal number, the blank controls injected subcutaneously with dimethyl sulfoxide plus normal saline, and the DES groups with DES at 0.1, 1, 10 and 100 microg/kg body weight, respectively, from embryonic day 9 (ED9) through ED17. Immunohistochemistry and RT-PCR were used to detect the expressions of LGR8 protein and mRNA in the gubernaculum testis of the ED18 fetuses and PND20 (postnatal day 20) offspring of the mice.</p><p><b>RESULTS</b>Histological analysis showed that the gubernaculum testis of the ED18 fetuses were well developed in both the normal and control groups, with an inner mesenchymal core and muscular outer layer. In contrast, the gubernaculum testis were poorly developed in the experimental groups, morphologically abnormal and without visible dividing line between the mesenchymal tissue and the muscular outer layer. No obvious differences were found in the gubernaculum testis development of the neonates between the normal and experimental groups. Positive immunostaining was seen in the mesenchymal core and muscular outer layer, but mainly in the latter. The expression of LGR8 was weaker in the experimental groups than in the normal group (P < 0.05), but that of LGR8 mRNA was increased in the high-dose (10 and 100 microg/kg) DES groups (P < 0.05). No obvious mutations were observed in the PCR products in any of the experimental groups.</p><p><b>CONCLUSION</b>Prenatal exposure to diethylstilbestrol affected the expression of LGR8 mRNA in the mouse gubernaculum testis, which suggests that diethylstilbestrol may induce cryptorchidism by interfering with the INSL3-LGR8 signaling system and consequently the development of the gubernaculum testis.</p>

Progress in researches on 46, XY disorders of sexual development / 中华男科学杂志

46, XY disorders of sexual development (46, XY DSD) are a group of complicated clinical conditions, which involve medical care, society, ethics and many other aspects. As chronic diseases, they necessitate long-term or even lifelong physical and mental follow-up and treatment. Early diagnosis and reasonable treatment could not only achieve appropriate development of the secondary sexual characteristics, but also effectively prevent gonadal malignancy. In recent years, environment pollution and other factors are contributing to the increasing incidence of 46, XY DSD all over the world. A deeper clinical insight into these disorders helps their earlier diagnosis and maximum improvement of prognosis. The etiology, new classification and treatment of 46, XY DSD are reviewed in this article.

Effects of diethylstilbestrol on cultured testis gubernacular cells in mice / 中华男科学杂志

<p><b>OBJECTIVE</b>To establish a primary culture of the testis gubernacular cells of Kunming mice, observe the morphological characteristics of the cells, and explore the effects of exogenous estrogens (EEs) on the development of the testis gubernacula in vitro.</p><p><b>METHODS</b>We removed the gubernacula from 3-day-old mice with the surgical magnifier and cultured the gubernacular cells. Then we detected the cell viability by trypan blue and cell morphology by HE staining. The subcultured cells were randomly divided into a blank control, a DMSO (0.1%, v/v) control, and 4 experimental groups (given 0.01, 0.10, 1.00 and 10.00 micdrog/ml of diethylstilbestrol [DES] dissolved in DMSO, respectively). After treated for 12, 24 and 48 hours, the gubernacular cells were observed for morphological changes and proliferation inhibition by CCK-8.</p><p><b>RESULTS</b>Most of the cultured gubernacular cells were fibroblasts, and a few were epithelioids. The primary cells showed a viability of 85%-90%. Dose- and time-dependent inhibition of cell proliferation was found in the four experimental groups at three different times, with statistically significant differences (P < 0.01).</p><p><b>CONCLUSION</b>Gubernacular cells can be cultured in vitro. EEs inhibit the proliferation of gubernacular cells in a dose- and time-dependent manner. An in- sight into the effects EES on cultured gubernacular cells is an effective approach to the study of their influence on the development of the reproductive system.</p>

A morphological study with serial histological slices on the normal and abnormal gubernacula in newborn male mice / 中华男科学杂志

<p><b>OBJECTIVE</b>To explore the feasibility of serial slices microscopic histological investigation for the elaborate evaluation of reproductive system malformations.</p><p><b>METHODS</b>Newborn male mice prenatally exposed to different doses of subcutaneously given diethylstilbestrol (DES) from gestational day 9 to 17 were treated by fixing parts of the abdomen in situ and setting them to transected serial slices. All the slices were stained, studied under the microscope and serially recorded by software. The gubernaculum was morphologically analyzed and its location and size were measured.</p><p><b>RESULTS</b>Morphologically, the gubernaculum could be identified clearly, its structure inhomogeneous from proximal to distal and dissymmetric from right to left. The environmental estrogen produced different effects on the morphology of the gubernaculum in different parts and most obviously affected its length.</p><p><b>CONCLUSION</b>Prenatal exposure to environmental estrogen has evident and general effects on the gubernacular development of newborn male mice. The morphological study with serial histological slices gives a precise and systematic evaluation of genital malformations.</p>

Relationship between insulin like hormone 3 and testicular descent and development / 中华男科学杂志

Testicular descent is an essential step in the course of reproductive system development. The mechanisms involved in the regulation of testis descent is not distinct. Gubernaculum has a very close relationship with testis descent. Maldescent of testis can cause abnormalities of genital system such as testicular underwent (cryptorchidism), dysplasia, tumor, infertility and low sexuality. Recently insulin like hormone 3 is a hotspot of concerning affecting gubernacular development and testicular descent. This article briefly reviews the advances in these aspects.

Biological characteristics of human umbilical cord-derived mesenchymal stem cells and their differentiation into neurocyte-like cells / 中华儿科杂志

<p><b>OBJECTIVE</b>To investigate the isolation and expansion of mesenchymal stem cells (MSCs) from human umbilical cord Wharton's jelly and their biological identities, and explore the possibility of inducing human umbilical cord-derived MSCs to differentiate into neurocyte-like cells.</p><p><b>METHODS</b>The growth and proliferative abilities of human umbilical cord-derived MSCs were observed, and their immunophenotypes were determined by flow cytometry. Salvia miltiorrhiza and beta-sulfhydryl alcohol were adopted to induce the cells to differentiate. The differentiated and undifferentiated cells were identified with immunocytochemistry. The pleiotrophin and nestin genes were measured by RT-PCR.</p><p><b>RESULTS</b>A population of human umbilical cord-derived MSCs were isolated from human umbilical Wharton's jelly; they were processed to obtain a fibroblast-like population of cells and could be maintained in vitro for extended periods with stable population doubling, and they were expanded as undifferentiated cells in culture for more than 10 passages, indicating their proliferative capacity. The human umbilical cord-derived MSCs were positive for CD(29), CD(44), CD(59), CD(105), but negative or weakly expressed the markers of hematopoietic cells such as CD(14), CD(33), CD(34), CD(28), CD(45) and CD(117). The important GVHD correlation markers were negative or weakly expressed, including CD(80) (B7-1), CD(86) (B7-2), CD(40) and CD(40L). Salvia miltiorrhiza beta-sulfhydryl alcohol could induce the MSCs to express nestin, a marker of neuronal precursor stem cells at early stage of differentiation. Later, they exhibited neural phenotypes, expressing beta-tubulin III and neurofilament (NF) and glial fibrillary acidic protein (GFAP). It was confirmed by RT-PCR that the MSCs could express pleiotrophin either before or after the induction of salvia miltiorrhiza, furthermore, after the induction the expression was markedly enhanced and the nestin gene was also expressed.</p><p><b>CONCLUSION</b>The human MSCs could be isolated from human umbilical cord Wharton's jelly, and it was easy to propagate these MSCs. The negative GVHD correlated markers might result from the fact that MSCs had no HLA barrier, which may suggest potential clinical significance. The MSCs are capable of differentiating into neurocyte-like cells and they may represent an alternative stem cell source for CNS cells transplantation.</p>

Advances in the endocrine factors affecting the development of gubernaculum testis / 中华男科学杂志

The testicular gubernaculum plays an important role in testicular descent and development. Its differentiation and development are affected by many factors. Androgens, calcitonin gene-related peptide (CGRP), insulin-like factor 3 (INSL3), Müllerian inhibiting substance (MIS), epidermal growth factor (EGF) and environmental estrogens (EEs) are involved in gubernacular development. The effect of CGRP, INSL3 and especially EEs on genital system has been attracted more attention.

Treatment of Disorders of Sex Development / 实用儿科临床杂志

Disorders of sex development(DSD) include anomalies of sex chromosomes,gonads,reproductive ducts,and genitalia.Here,the term "intersex" is avoided because of its imprecision.The purpose of this review is to assist health care professionals in the provision of treatment,education,and support to children born with DSD and to their families.The goal of DSD treatment is to achieve the long-term physical,psychological,and sexual well-being of the patients.In the case of DSD it involves several principles.Providing medical and surgical care to deal with a complication that threatens to the patient's physical well-being;minimizing the potential for the patient and family to feel ashamed,stigmatized,or overly obsessed with genital appearance;delaying elective surgical and hormonal treatments until the patient can actively participate in decision-making;telling the truth to the family and the child and addressing psychosocial distress of the children and their parents.This approach is termed as "patient-centered".

Human Umbilical Cord Blood Mesenchymal Stem Cells Colonizing in Hypoxic-Ischemic Encephalopathy Rats Brain / 实用儿科临床杂志

Objective To explore human umbilical cord blood mesenchymal stem cells(MSCs)colonization in hypoxic-ischemic encephalopathy(HIE)rats brain.Methods Models of 7-day-old newborn rats with HIE brain injury were established.Meanwhile,on the same day,MSCs were transplanted with Hoechst 33258 for 24 hours into rats models marked by flurescent nucleotide dye injected through caudal vein or with stereotactic instrument.After 15-30 days,then MSCs were detected with fluorescene microscope.Results With the improved rice methods,HIE animal model was successfully attained.Majority of MSCs were distributed in the cortex,hippocampus of the lesioned hemisphere,especially in the forehead.And abtained a good fusion with HIE rats brain tissue.Conclusion Human umbilical cord blood MSCs can be cultured,when transplanted into the HIE rats model,they can move into intracranial,and integer with rats brain tissue.

Human umbilical cord Wharton's Jelly-derived mesenchymal stem cells differentiation into nerve-like cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>The two most basic properties of mesenchymal stem cells (MSCs) are the capacities to self-renew indefinitely and differentiate into multiple cells and tissue types. The cells from human umbilical cord Wharton's Jelly have properties of MSCs and represent a rich source of primitive cells. This study was conducted to explore the possibility of inducing human umbilical cord Wharton's Jelly-derived MSCs to differentiate into nerve-like cells.</p><p><b>METHODS</b>MSCs were cultured from the Wharton's Jelly taken from human umbilical cord of babies delivered after full-term normal labor. Salvia miltiorrhiza and beta-mercaptoethanol were used to induce the human umbilical cord-derived MSCs to differentiate. The expression of neural protein markers was shown by immunocytochemistry. The induction process was monitored by phase contrast microscopy, electron microscopy (EM), and laser scanning confocal microscopy (LSCM). The pleiotrophin and nestin genes were measured by reverse transcription-polymerase chain reaction (RT-PCR).</p><p><b>RESULTS</b>MSCs in the Wharton's Jelly were easily attainable and could be maintained and expanded in culture. They were positive for markers of MSCs, but negative for markers of hematopoietic cells and graft-versus-host disease (GVHD)-related cells. Treatment with Salvia miltiorrhiza caused Wharton's Jelly cells to undergo profound morphological changes. The induced MSCs developed rounded cell bodies with multiple neurite-like extensions. Eventually they developed processes that formed networks reminiscent of primary cultures of neurons. Salvia miltiorrhiza and beta-mercaptoethanol also induced MSCs to express nestin, beta-tubulinIII, neurofilament (NF) and glial fibrillary acidic protein (GFAP). It was confirmed by RT-PCR that MSCs could express pleiotrophin both before and after induction by Salvia miltiorrhiza. The expression was markedly enhanced after induction and the nestin gene was also expressed.</p><p><b>CONCLUSIONS</b>MSCs could be isolated from human umbilical cord Wharton's Jelly. They were capable of differentiating into nerve-like cells using Salvia miltiorrhiza or beta-mercaptoethanol. The induced MSCs not only underwent morphologic changes, but also expressed the neuron-related genes and neuronal cell markers. They may represent an alternative source of stem cells for central nervous system cell transplantation.</p>

Expression of glucocorticoid receptor isoforms in cutaneous hemangiomas and vascular malformations / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Hemangiomas are the most common tumors in children. Some hemangiomas may require intervention because of their location, size, behavior, or potential for important complications. Pharmacological therapy with glucocorticoids is the mainstay treatment, but there is no consensus on therapeutic regimens or candidate selection, therapeutic efficacy varies, and the mechanism mediating the beneficial effects of glucocorticoids remains unclear. This study was performed to investigate the expression patterns of the glucocorticoid receptor (GR) and its alpha isoform (GRalpha) in cutaneous hemangiomas and vascular malformations.</p><p><b>METHODS</b>SP immunohistochemical technique was used to examine the expression of GR(e-20) (GR) and GR(p-20) (GRalpha) on vascular endothelial cells in 80 specimens that included 33 proliferating hemangiomas, 32 involuting hemangiomas, 7 vascular malformations as well as 8 normal skin tissues, all obtained from infants and children. GR and GRalpha expression in prepared tissue slides were examined using automated computer-assisted microscopic analysis. Mean gray scale values were compared among the various tumor types.</p><p><b>RESULTS</b>The mean gray scale values of GR were 127.0 +/- 6.4 and 121.4 +/- 6.6 in hemangiomas and vascular malformations respectively, but this difference was not statistically significant (P = 0.104). However, these values were all markedly higher than that of normal skin, which was only 108.6 +/- 6.8 (P = 0.001 and P = 0.000 for comparison with hemangiomas and vascular malformations respectively). The gray scale of GR in proliferation and involuting hemangiomas were 127.9 +/- 4.8 and 126.0 +/- 5.8 respectively, but this difference was not significant (P = 0.146). However, GRalpha expression in hemangiomas, vascular malformations and normal skin declined gradually in stepwise fashion (127.3 +/- 5.4, 120.4 +/- 6.1 and 109.9 +/- 5.3 respectively; P < 0.001). GRalpha expression was higher in proliferating hemangiomas than in involuting hemangiomas (127.2 +/- 6.3 and 122.5 +/- 6.3; P = 0.004).</p><p><b>CONCLUSIONS</b>GR and GRalpha are strongly expressed in hemangiomas and vascular malformations. The expression of GRalpha is closely related to the phase of the hemangioma. Determination of GR and GRalpha may be a positive significance to understand the information of hemangiomas and vascular malformations and may further help determining proper strategies of steroid therapy for hemangiomas and vascular malformations.</p>

Diagnosis,Treatment and Prognosis of Neonatal Intestinal Atresia / 实用儿科临床杂志

Objective To explore diagnosis,treatment and prognosis of intestinal atresia(IA) in neonates,so as to increase the healing rate.Methods Surgical treatment was given to 29 cases of congenital IA from Apr.2004 to Nov.2008.Comprehensive analysis was carried out on clinical manifestations and flat X-ray.All cases were with vomiting,15 cases were with normal fetal faeces.Stomach and intestinal atresia were observed in the upper abdomen in 19 cases,fluid levels in generous stomach and expended intestines were seen in 21 cases by X-ray.All children underwent operation,including ileum atresia(16 cases),duodenal atresia(7 cases),and jejunal atresia(6 cases).The types of atresia were type Ⅰ(9 cases),type Ⅱ(9 cases),type Ⅲa(9 cases),type Ⅲb(1 case) and type Ⅳ(1 case).Nine cases were treated with septum resection and then anastomosed longitudinally,3 cases underwent end-to-end anastomose and 17 underwent end-to-back anastomose.All cases were served in intravenous hyperalimentation.Results All cases lived postoperatively,and 1 child gave up for postoperatively treatment.Twenty-eight cases were cured.There was 1 case of obstruction of anastomatic,and was cured by non-operatively treatment.There was no anastomatic leakage and wound disruption.Twenty-six cases were followed up for 3 months and up to 4 years.These symptoms had no relapse.Conclusion Early diagnosis,appropriate surgical procedure and surrounding operation management are crucial for improving the cure rate.

Transcatheter arterial chemoembolization and delayed surgery for infant hepatoblastoma / 实用儿科临床杂志

Objective To evaluate the effect of transcatheter arterial chemoembolization (TACE) and delayed surgery for infant hepatoblastoma.Methods TACE was performed with the initial digital subtractive angiography (DSA) under general anesthesia 1-3 times in 8 infants with huge hepatoblastoma, whose age was 2 to 12 months. DSA was done via arterials in hepatoblastoma each time before chemoembolization. The arterials were perfused with chemodrugs and suspensions in ultrasome iodized oil , and were blocked with spring rings. DSA findings indicated that the tumor shrank without new tumorous arterials after 1 month in 6 cases, and 4 of them showed no tumorous staining, and the delayed surgery was performed successfully 1 week later in 6 infants. One boy underwent systemic chemotherapy alone during 6 months after 3 times of TACE. Results TACE therapy did not encounter any major technical problem or toxic reaction caused by chemotherapy. The following DSA test 4 weeks later did not detect any new tumorous vessels in 6 cases. Six children received TACE and surgery had been followed-up with no tumor recurrence for months averagely. The boy underwent TACE and venous chemotherapy for 6 months , without surgery , had been followed-up for 48 months until the present report. CT, AFP and DSA did not show any hints of tumor recurrence. Six cases receiving 3 times TACE combined with surgery survived without tumor recurrence. Conclusions TACE is a very effective, safe and helpful therapy for hepatoblastoma, which stressed the repeated use of spring ring to block tumor vessels lastingly if necessary. If surgery is required, DSA test is needed beforehand to detect new tumorous vessels or neoplasm. If there is any , TACE is repeated. TACE combined with surgery may provide an additional promising choice in the treatment of hepatoblastoma, and repeated TACE alone may cure hepatoblastoma in infants.

Effect of prenatal exposure to diethylstilbestrol on gubernacular development in fetal male mice / 亚洲男科学杂志(英文版)

<p><b>AIM</b>To study the effect of prenatal exposure to diethylstilbestrol (DES) and the role of actin and proliferating cell nuclear antigen (PCNA) on testicular gubernaculum development in fetal male Kunming mice.</p><p><b>METHODS</b>Pregnant mice were randomly assigned to 6 groups and injected with DES subcutaneously from gestational day 9 (E9) to day 17 (E17) at doses of 0, 25, 50, 100, 200 microg.kg-1.d-1 in 0.2 mL dimethyl sulfoxide (DMSO). On E17 they were sacrificed and fetuses quickly removed for fixation. Male fetuses were sliced on serial coronal plane. Histological changes were observed under the light microscope (LM) and ultrastructural changes with the scanning and transmission electron microscopes (SEM and TEM). The expression intensity of actin and PCNA in the gubernacula was quantitated by immunohistochemistry.</p><p><b>RESULTS</b>The mortality of the fetuses was higher in the DES-treated groups than that in the DMSO and saline controls (P<0.05). Under LM the gubernacula were seen to be poorly developed with smaller bulbs. On SEM the bulbs lose the clear demarcation between the mesenchymal inner core and the muscular outer layer and looked like a small cone instead of the normal cylindrical appearance. On TEM there were some smaller disordered myofibrils and sparse cytoplasmic organelles in the gubernacular muscular cells of the treated groups. The expression intensity of actin and PCNA in the gubernacula was significantly weaker in the treated groups than that in the DMSO and saline controls (P<0.05).</p><p><b>CONCLUSION</b>DES induces underdevelopment of the gubernacula in a dose-dependent manner in fetal male mice and down regulates the actin and PCNA expression.</p>