[The role of bronchoscopy in slide tracheoplasty in children]. / 支气管镜在儿童Slide气管成形术中的应用.

OBJECTIVES: To study the role of bronchoscopy in slide tracheoplasty. METHODS: A retrospective analysis was conducted on the diagnosis and treatment of four children with tracheal stenosis admitted to Hunan Provincial People's Hospital from 2017 to 2020. The role of bronchoscopy was summarized in the preoperative evaluation, intraoperative positioning and measurement, and postoperative wound evaluation and treatment during slide tracheoplasty. RESULTS: Bronchoscopy evaluation before slide tracheoplasty showed that 3 of the 4 children had complete trachea rings, 2 had pulmonary artery sling, and 2 had multiple stenosis. Slide tracheoplasty was performed in the hospital on 3 children, and the midpoint of the stenosis segment was judged under bronchoscopy, and the length of the stenosis segment was measured, which assisted in the resection of the stenosis segment of the trachea. The pathogens were identified by lavage after the surgery. One child who developed scar traction 9 months after slide tracheoplasty in another hospital was improved by interventional treatment under bronchoscopy. Mucosal changes were found under bronchoscopy in 2 children 4 days after surgery, and the treatment plan was adjusted. One month after surgery, 2 children had granulation hyperplasia, which was improved by cryotherapy under bronchoscopy. One child abandoned treatment due to anastomotic necrosis and died. Three survivors were followed up for over 6 months with good prognosis, but all had tracheobronchial malacia. CONCLUSIONS: Bronchoscopy can be used for the management of slide tracheoplasty in children with tracheal stenosis, which is helpful to postoperative rehabilitation and follow-up.

[Clinical effect of propranolol in the treatment of respiratory hemangioma in infants and young children].

OBJECTIVE: To study the clinical effect of oral propranolol in the treatment of respiratory hemangioma in infants and young children. METHODS: A retrospective analysis was performed from the chart review data of children with respiratory hemangioma treated by oral propranolol and diagnosed by bronchoscopy and laryngeal plain enhanced CT/MRI from November 2012 to December 2019. RESULTS: A total of 20 children were enrolled. All children had improvement in the symptoms of laryngeal stridor and dyspnea after oral administration of propranolol for 1-2 days. The median treatment time was 10 months (range 6-12 months). The median follow-up time was 10 months (range 3-15 months). Of the 20 children, 19 (95%) achieved regression of tumor, and 1 (5%) experienced an increase in tumor size during reexamination at 6 months after drug withdrawal and had no recurrence after the treatment with an increased dose of propranolol for 6 months. Only 1 child (5%) had adverse reactions, and 1 child (5%) was still under treatment. CONCLUSIONS: Oral propranolol can quickly relieve the symptoms such as dyspnea and achieve tumor regression, with few adverse events, and it is therefore an effective method for the treatment of respiratory hemangioma in infants and young children.

[Relationship between viral load of human bocavirus and clinical characteristics in children with acute lower respiratory tract infection].

OBJECTIVE: To investigate the prevalence of human bocavirus (HBoV) in children with acute lower respiratory tract infection and to explore the relationship between the viral load of HBoV and the clinical characteristics of acute lower respiratory tract infection in children. METHODS: A total of 1 554 nasopharyngeal aspirates from children who were hospitalized due to acute lower respiratory tract infection between March 2011 and March 2014 were collected. Quantitative real-time PCR was used to detect 12 RNA and 2 DNA viruses, adenovirus (ADV) and HBoV, and to measure the viral load of HBoV in HBoV-positive children. A comprehensive analysis was performed with reference to clinical symptoms and indicators. RESULTS: In the 1 554 specimens, 1 212 (77.99%) were positive for viruses, and 275 (17.70%) were HBoV-positive. In HBoV-positive cases, 94.9% were aged <3 years, and there were more males than females. In the 275 HBoV-positive cases, 45 (16.36%) had single infection, and 230 (83.64%) had mixed infection. There was no significant difference in viral load between children with single infection and mixed infection (P>0.05). The patients with fever had a significantly higher viral load than those without fever (P<0.05). The children with wheezing had a significantly higher viral load than those without wheezing (P<0.05). There was no significant difference in viral load between children with mild, moderate, and severe acute lower respiratory tract infection (P>0.05). CONCLUSIONS: HBoV is one of the important pathogens of acute lower respiratory tract infection in children. Children with a higher viral load of HBoV are more likely to experience symptoms such as fever and wheezing. However, the severity of disease and mixed infection are not significantly related to viral load.

Influence of blastocysts morphological score on pregnancy outcomes in frozen-thawed blastocyst transfers: a retrospective study of 741 cycles.

The influence of inner cell mass (ICM) and trophectoderm (TE) score on pregnancy outcomes in frozen-thawed blastocyst transfer cycles was analyzed. A retrospective analysis of 741 cycles of frozen-thawed blastosysts transfer was performed. All cycles were divided into four groups based on the number and morphological score of blastocysts: S-ICM B/TE B group (n=91), the single blastocyst transfer of ICM B and TE B; D-ICM B/TE B group (n=579), double blastocysts transfer of ICM B/TE B; D-ICM B/TE C group (n=35), double blastocysts transfer of ICM B/TE C; and D-ICM C/TE B group (n=36), double blastocysts transfer of TE B/ICM C. The pregnancy outcomes were compared among the four groups. As compared with D-ICM B/TE C group, the clinical pregnancy rate, implantation rate and multiple pregnancy rate were increased in D-ICM B/TE B group (74.96% vs. 57.14%, 57.43% vs. 37.14%, and 48.62% vs. 25%, respectively, P<0.05 for all). Clinical pregnancy rate and implantation rate in D-ICM B/TE B group were also higher than in D-ICM C/TE B group (74.96% vs. 50%, and 57.43% vs. 33.33%, both P<0.05). Multivariable Logistic regression analysis indicated that ICM score was a better predictive parameter for clinical pregnancy (OR=3.05, CI 1.70-5.46, P<0.001), while the trophectoderm score was a better one for early abortion (OR=0.074, CI 0.03-0.19, P<0.001). Clinical pregnancy rate and multiple pregnancy rate in S-ICM B/TE B group were significantly lower than those in D-ICM B/TE B group (46.15% vs. 74.96%, and 2.38% vs. 48.62%, both P<0.05), but there was no significant difference in the implantation rate between the two groups. It was suggested that the higher score of ICM and TE may be indicative of the better pregnancy outcomes. The ICM score is a better predictor of clinical pregnancy than TE, while TE score is a better one in predicting early abortion. Single ICM B/TE B blastocyst transfer in frozen-thawed cycles can also get satisfactory pregnancy outcomes.

[Clinical features and pathogens of plastic bronchitis in children: an analysis of 9 cases].

OBJECTIVE: To study the clinical features and pathogens of plastic bronchitis in children. METHODS: A retrospective analysis was performed on the clinical data of 9 children who were diagnosed with plastic bronchitis between January 2011 and December 2012. RESULTS: Plastic bronchitis began with a fever and cough in all cases, followed by progressive dyspnea on days 1-3 of onset; unilateral or bilateral decreased breath sounds and hepatosplenomegaly were found; complications included respiratory failure (6 cases), toxic encephalopathy (6 cases), toxic hepatitis (7 cases), shock (3 cases), heart failure (3 cases), and renal failure (2 cases). Chest X-ray or chest CT showed single and multiple lobar or segmental consolidation and atelectasis, as well as pleural effusion (4 cases). The bronchofibroscopy revealed some grey-white mucus plugs that blocked bronchial openings and aspirates of bronchial shape. Influenza viruses (IFVs) were detected in all cases, including IFV-A (6 cases, 67%) and IFV-B (3 cases, 33%). Mixed infection with IFV-A and Mycoplasma pneumoniae (MP)/bacteria was found in 50% of all cases. In the three cases of IFV-B infection, one was complicated by MP infection. Nine patients were given treatment of antibiotics, hormones, gamma globulin and necessary respiratory support, and also were given removal of endogenous foreign body by bronchoscopy. Five patients were given antiviral therapy of oseltamivir. Seven cases cured, and 2 died. CONCLUSIONS: Plastic bronchitis and severe pneumonia are similar in clinical manifestations. IFVs are the main pathogen. In addition to anti-infection treatment, hormone, gamma globulin, respiratory support, and other conventional treatments, endogenous foreign body removal by bronchofibroscopy and early antiviral therapy with oseltamivir have good efficacy.

Prevalence of human metapneumovirus in children with acute lower respiratory infection in Changsha, China.

Human metapneumovirus (hMPV) causes acute respiratory infections in children. The prevalence and clinical characteristics of hMPV were determined in nasopharyngeal aspirates of children in Changsha, China. Reverse transcription-polymerase chain reaction (RT-PCR) or PCR was employed to screen for both hMPV and other common respiratory viruses in 1,165 nasopharyngeal aspirate specimens collected from children with lower respiratory tract infections from September 2007 to August 2008. All PCR products were sequenced, and demographic and clinical data were collected from all patients. Seventy-six of 1,165 (6.5%) specimens were positive for hMPV, of which 85.5% (65/76) occurred in the winter and spring seasons. The hMPV coinfection rate was 57.9% (44/76), and human bocavirus was the most common virus detected in conjunction with hMPV. Phylogenetic analysis revealed that 94.7% of the hMPV detected were of subgroup A2, 5.3% were subgroup B2, and none belonged to either the A1 or B1 subgroups. No significant differences were found in terms of the frequency of diagnosis and clinical signs between either the co- and mono-infection groups, or between patients with and without underlying diseases. It was concluded that hMPV is an important viral pathogen in pediatric patients with lower respiratory tract infections in Changsha. Only hMPV genotypes A2 and B2 were co-circulating in this locality; human bocavirus was the most common coinfecting virus, and coinfection did not affect disease severity.

[Status of enterovirus infection in children with acute lower respiratory tract infection].

OBJECTIVE: To investigate the status of enterovirus (EV) infection in children with acute lower respiratory tract infection (ALRTI). METHODS: A total of 404 samples (with odd numbers) of nasopharyngeal aspirates were collected from the children who were hospitalized in the Children's Medical Center, Hunan Provincial People's Hospital due to ALRTI between September 2007 and April 2008. The conserved sequence in the 5'-noncoding region of EV was used to design the primer, and nested RT-PCR was performed to detect EV in the samples. RESULTS: Of the 404 samples, 19 (4.7%) were EV-positive, and mostly taken from children under 3 years of age (95%); there was no significant difference in the detection rate between male and female children. Of the EV-positive children, 13 (68%) were clinically diagnosed with bronchial pneumonia, and 6 (32%) with bronchiolitis; 90% of them showed symptoms of fever, 84% had a cough, 63% had asthma, and 63% had complications mainly including diarrhea (6 cases), granulocytopenia (4 cases), and acute respiratory distress syndrome (2 cases). In addition, 26% of the EV-positive children had leukocyte disorder, more than half had liver dysfunction, and a few had myocardial involvement. CONCLUSIONS: EV is a pathogen that should not be neglected in children with ALRTI. For these children, close attention should be paid to the epidemiological status and clinical features of EV infection, and blood routine examination, liver function test and myocardial enzyme assay should be carried out periodically to improve prognosis.

[Viral etiology and risk factors for severe community-acquired pneumonia in children].

OBJECTIVE: To study the virus spectrum of severe community-acquired pneumonia (CAP) and risk factors for the disease in children. METHODS: Respiratory secretion specimens were collected from 1096 children hospitalized with CAP from June 2007 to November 2008, including 100 cases of severe CAP. Respiratory viruses were detected by PCR, nest-PCR or RT-PCR. Clinical data on the children were analyzed by univariate and multivariate logistic regression analysis for examining risk factors for severe CAP. RESULTS: Viral pathogens were isolated from 82 (82%) of the 100 cases with severe CAP. RSV was the most common (37%), followed by HBoV (25%) and HRV (18%). Mixed infection was noted in 32 cases (32%). The presence of underlying diseases (OR=6.623, P<0.01) and RSV infection (OR=1.672, P<0.05) were risk factors for severe CAP in children, while age was a protective factor (OR=0.475, P<0.01). CONCLUSIONS: RSV is the most frequent viral pathogen in children with severe CAP. The presence of underlying diseases and RSV infection may be risk factors for severe CAP, while age is a protective factor.

[Viral etiology of 1165 hospitalized children with acute lower respiratory tract infection].

OBJECTIVE: To explore the viral etiology of acute low respiratory tract infection (ALRTI) among hospitalized children in Changsha of Hunan Province of China. METHODS: Nasopharyngeal aspirates were collected from 1165 hospitalized children with ALRTI in Changsha from September 2007 to August 2008. Respiratory syncytin virus (RSV), human rhinovirus (HRV), influenza virus A (IFVA), influenza virus B (IFVB), parainfluenza 1-3 (PIV 1-3), human metapneumovirus (hMPV), human coronaviruses NL63 (HCoV-NL63), and human coronaviruses HKU1 (HCoV-HKU1) were detected by reverse transcription polymerase chain reaction (RT-PCR). Adenovirus (ADV) and human bocavirus (HBoV) were detected by standard polymerase chain reaction (PCR). WU polyomaviruses (WUPyV) and KI polyomaviruses(KIPyV) were detected by nested PCR. The positive samples further underwent genetic sequencing. RESULTS: Among the 1165 nasopharyngeal aspirates, viruses were detected in 871 samples (74.76%), among which RSV (27.03%) was the most common virus, followed by HRV (17.33%), PIV3 (13.73%), HBoV (8.67%) and hMPV (6.52%). The overall positive rate of viral detection showed no significant differences between males and females (X2=2.241, P=0.134), whereas the positive rates of PIV3, hMPV, and HBoV in males were higher than in females. The positive rate of viral detection showed significant differences among different age groups (X2=10.934, P=0.027), and the highest positive rate was noted in the age group of 6 months to 1 year. Furthermore, the overall positive rate of viral detection showed a significant difference in term of seasonal distribution, with a peak prevalence in winter. CONCLUSIONS: Virues predominate in the etiology of pediatric ALRTI in Changsha, and RSV, HRV and PIV3 are the main viruses for ALRTI. HBoV and hMPV have become increasingly important. Viral infection-associated ALRTI shows a prevail in the age group of 6 months to 1 year as well as in winter.

[Protective effects of lidocaine on hyperoxia-exposed type â ¡alveolar epithelial cells from premature rats].

OBJECTIVE: To investigate the effects of lidocaine on apoptosis and proliferation of hyperoxia-exposed type Ⅱalveolar epithelial cells (AECⅡ) from premature rats. METHODS: Primary cultured AEC Ⅱ isolated from premature rats were randomly divided into four groups: air, air+ lidocaine (20 µg/mL), 95%O2/5%CO2, and 95%O2/5%CO2+ lidocaine. The cells in each group were collected 24 hrs after culture in ordinary incubators (37℃,5%CO2). The proliferation and apoptosis of AEC Ⅱ were detected by flow cytometry. Protein levels of proliferating cell nuclear antigen (PCNA) were measured by Western blot. RESULTS: The apoptosis rate of AECⅡincreased, the percentages of G2/M and S phase cells decreased and the protein levels of PCNA decreased significantly in the group exposed to 95%O2/5%CO2 compared with the group exposed to air (P<0.01). Lidocaine treatment decreased apoptosis rate of AECⅡ, increased percentage of G2/M and S phase cells, and increased protein levels of PCNA. CONCLUSIONS: Hyperoxia can increase apoptosis and inhibit proliferation of AECⅡ in premature rats. Lidocaine may have protective effects against the AECⅡ injuries.

[Modified transverse tibial bone transplantation and microcirculation reconstruction for the treatment of end-stage diabetic foot].

OBJECTIVE: To explore clinical effect of modified transverse tibial bone transfer microcirculation reconstruction in treating end-stage diabetic foot. METHODS: From August 2016 to June 2018, 87 patients with diabetic foot treated with modified tibial transverse bone removal and microcirculation reconstruction, inclduing 54 males and 33 females;aged from 39 to 95 years old with an average of (68.9±11.3) years old;2 patients were grade 2, 37 patients were grade 3 and 50 patients were grade 4 according to Wagner's classification;the courses of diabetic were for 10 to 16 years with an average of (13.0±2.2) years;the courses of diabetic feet were for 21 to 48 days with an avergae of (34.2±8.6) days. Postoperative comlications were observed. Skin temperature, visual analogue scale(VAS) and ankle brachial index(ABI) and wound healing were recorded before and 3 months after operation. RESULTS: All patients were followed up for 4 to 19 months with an average of (12.6±2.8) months. Two patients occurred subcutaneous tissue liquefaction and seepage under needle passage during bone transfer, and scabed without special treatment. One patient was performed amputation above 5 cm of ankle joint because of severe infection, and 1 patient occurred re-ulceration at 1 year after wound healing, bone transfer was performed again at the same site, and was completely healed at 8 weeks after operation. The healing time of wound ranged from 3 to 24 weeks with an average of (11.9± 3.8) weeks. Foot skin temperature before operation was (28.9±0.91) ℃, and increased to (31.70±0.32)℃ at 3 months after operation(t=5.72 P=0.006);VAS score before opertaion was (7.80±0.72), and improved to (2.20±0.13) at 3 months after operation (t=25.38, P=0.000);ABI beforeoperation was(0.48±0.30), increased to(0.98±0.24) at 3 months after oeprtaion(t= 14.68, P=0.000). CONCLUSION: Modified lateral tibial bone transfer could effectively reconstruct microvascular network under lower leg, promote recovery of peripheral blood vessels, and promote wound healing of foot, reduce or avoid amputation. At the same time, the improved osteotomy is one of the effective methods for the treatment of diabetic foot which has advantags of less trauma, simple opertaion.

[The effect of the split of sperm nuclei on the yield of human sperm RNA].

OBJECTIVE: To study the effect of the split of sperm nuclei on the yield of RNA from human sperm. METHODS: Human sperm were purified by two sequential centrifugations through 40:80 discontinuous gradients of Percoll. Human leukocytes separated from peripheral blood were used as the control. Total RNAs from purified sperm and leukocytes were extracted with both TRIzol and RNeasy Kit. The RNAs from equal number of cells were reverse-transcribed, and quantified by the levels of beta-ACTIN mRNA, which were evaluated by real time polymerase chain reaction. RESULTS: TRIzol failed to digest majority of sperm nuclei even the incubation time was prolonged to 1 h, while no sperm nucleus was found under the light microscope after 1 min digestion with RLT buffer of the RNeasy Kit. Both reagents can digest the nuclei of human leukocytes well. The amount of RNA per 10(6) sperms isolated with RNeasy Kit (149.8+/-24.5 ng) was 4-fold higher (P=0.01) than that extracted with TRIzol (35.5+/-4.0 ng per 10(6) spermatozoa; n=3). The similar yields of the leukocyte RNAs extracted with RNeasy Kit and TRIzol [(765.5+/-229.8) and (958.8+/-201.0) ng per 10(6) cells respectively; n=3, P=0.168] excluded the possibility of different efficacy of these two reagents in RNA isolation. CONCLUSION: The split of sperm nuclei is important to the yield of RNA in the human sperm RNA extraction. The nucleus may be the major area for human sperm RNA repositories.

[Y chromosome microdeletions of 664 Chinese men with azoospermia or severe oligozoospermia].

OBJECTIVE: To explore the incidence and location of Y chromosome microdeletions in Chinese azoospermia and severe oligozoospermia, as well as the relationship between the deletion region and testicular phenotype. METHODS: Semen samples or blood samples were collected from 664 Chinese patients (584 with azoospermia and 80 with severe oligozoospermia). DNA was extracted by incubating cells with a lysis buffer containing polymerase chain reaction (PCR) buffer and proteinase K, and was assayed for deletion of 15 sequence tagged sites (including 6 loci recommended by European Academy of Andrology and European Molecular Genetics Quality Network (EAA/EMQN) distributed in AZFa, AZFb and AZFc by 4 multiplex PCRs. The histological phenotypes of testes of some azoospermic patients harboring Y chromosome microdeletion were studied by fine needle aspiration. RESULTS: Sixty-six (11.3%) cases of microdeletions were found in the 584 patients with azoospermia, and deletions of AZFc region are the leading group (72.7% of all deletions), followed by AZFbc (13.6%), AZFabc (6.1%), AZFb (4.5%) and AZFa (3.0%). In the 80 men with severe oligozoospermia, 10 (12.5%) cases of AZFc microdeletions were detected. While azoospermia (n=19) with AZFc region deletion showed variable testicular phenotype, deletions of AZFb+c and AZFa+b+c (n=7) resulted in severe impaired spermatogenesis characterized by Sertoli cell only syndrome and spermatogenic arrest at spermatogonia. CONCLUSION: In the Chinese men with azoospermia and severe oligozoospermia, the incidence of Y chromosome microdeletions and the frequency of the deletions of the three AZF regions are similar to those described previously in other populations. Massive deletions of AZFb+c and AZFa+b+c impair spermatogenesis severely.

[Detection of Y chromosome microdeletions in semen of patients with azoospermia: study of 241 cases].

OBJECTIVE: To set up a simple and reliable method to screen Y chromosome microdeletions in semen of azoospermic patients, and to explore the incidence and loci of Y chromosome microdeletions in Chinese azoospermia. METHODS: Two hundred and forty-one semen samples, 51 containing blood, were collected from 241 Chinese azoospermic patients. 45 normal semen samples and 1 anticoagulated blood sample from female were used as controls. DNA was quickly abstracted by incubating the cells with a lysis buffer containing polymerase chain reaction (PCR) buffer and protease K, and was used to detect the deletion of 15 kinds of sequence tagged site (STS) distributed in AZFa, AZFb, and AZFc by 4 sets of multiplex PCR. Agarose electrophoresis was used to observe and compare the PCR products from the semen samples and their corresponding blood samples, and the PCR products from 1 semen sample were confirmed by sequencing. RESULTS: All multiplex PCR reactions were amplified successfully. The sequencing of the PCR products from the semen samples confirmed the PCR reactions. No microdeletion was detected in the 45 normal semen samples. Microdeletion was found in 26 out of the 241 semen samples of azoospermic patients (10.8%): 2 patients (7.7%) had the deletions located in AZFa, 2 patients (7.7%) in AZEb, 3 patients (11.5%) in both AZFb + AZFc, and other 19 patients (73.1%) in AZFc. The detection results of the blood samples were completely consistent with those of the semen samples. The STS deletion recommended by European Academy of Andrology (EAA) and European Molecular Genetics Quality Network (EMQN) were all found in these 26 cases. CONCLUSIONS: The semen samples of azoospermic patients present a convenient, reliable, and noninvasive substitute for blood in screening of Y chromosome microdeletions, and can be employed in study and clinical examination. The EAA/EMQN recommendations allow the detection of complete AZF deletions in Chinese azoospermia.

[A novel class of testis-specific small RNAs: piRNAs].

Eukaryotes produce various types of 19-30 nt small RNAs, which act as guides to the regulation of gene expressions, such as mRNA degradation and translational repression. The Argonaute family members related to small RNA functions fall into 2 subfamilies. One is the AGO subfamily, whose 4 members distribute widely, confirmedly bind to miRNAs and siRNAs and inhibit the expression of target mRNAs through a pathway like RNA interference. The other is the PIWI subfamily, including PIWI, Aubgine (AUB) and AGO3, exclusively expressed in the testis. Recently, four research groups have isolated a new class of small RNAs from the mammalian testis, which interacts with the PIWI subfamily, hence named piwi-interfering RNAs (piRNAs), and is suggestive of an important role in spermatogenesis.

Inhibition of the urokinase-type plasminogen activator by triplex-forming oligonucleotides in rat Sertoli cells: a new contraceptive alternative?

Urokinase-type plasminogen activator (uPA), expressed in Sertoli cells in the testis, is closely related with tight junctions of blood-testis barrier (BTB), and it has been considered as a potential contraceptive target. In the present study, the antigene effects of triplex-forming oligodeoxynucleotides (TFO) targeting uPA in rat Sertoli cells were investigated in vitro. The stable triplexes, formed by uPA specific TFOs under physiological conditions, were tested by means of electrophoretic mobility shift assays (EMSA). Although tPA, another form of plasminogen activators (PAs), partially compensated the lose of PAs activities, uPA mRNA and protein were significantly reduced as demonstrated by real-time reverse transcription PCR and a chromogenic assay, after the treatment of Sertoli cells with uPA specific TFOs at a concentration of 330 nM. The capacity of TFOs resistance to nuclease degradation was enhanced by the phosphorothioated on the backbone of the oligonucleotides. Our results indicated that the TFOs can downregulate uPA expression and uPA might be an alternative contraceptive target.

Gene expression changes of urokinase plasminogen activator and urokinase receptor in rat testes at postnatal stages.

AIM: To investigate the gene expression changes of urokinase plasminogen activator (uPA)/urokinase receptor (uPAR) in rat testes at postnatal stages and explore the effects of uPA/uPAR system on the rat spermatogenesis. METHODS: The mRNAs of uPA and uPAR in rat testes were measured by using real-time quantitative polymerase chain reaction (PCR) at postnatal days 0, 5, 10, 15, 21, 28, 35, 42, 49 and 56, respectively. RESULTS: The tendencies of uPA and uPAR mRNA expression were similar at most postnatal stages except for D(0). The expression of uPAR mRNA in rats testes was relatively higher than that of uPA at postnatal D(0), and both were decreased until D(21), increased obviously at postnatal D(28), reached a peak at postnatal D(35), then declined sharply at postnatal D(42) and retained at a low level afterwards. CONCLUSION: The uPA/uPAR system may be strongly linked to spermiation and spermatogenesis via regulating germ cell migration and proliferation, as well as promoting the spermiation and detached residual bodies from the mature spermatids.

[Effect of urokinase-type plasminogen activator on the mitochondrial membrane potential of mouse capacitated-spermatozoa in vitro].

OBJECTIVE: To study the mechanism of uPA improving sperm capacitation by investigating the effect of uPA on the mitochondrial function of mouse capacitated-sperm in vitro. METHODS: Mitochondrial function of mouse capacitated-spermatozoa was evaluated through the assessment of mitochondrial membrane potential using JC-1 performed by flow cytometer and fluorescent microscope respectively. The experiment and the control groups were designed according to the presence or absence of uPA, each divided into 5 subgroups based on the different time of uPA treatment (or BWW in the control groups) at 0, 5, 15, 30 and 60 min respectively. RESULTS: (1) Compared with that at 0 min, the mean fluorescence intensity of JC-1 within the spermatozoal body and the percentage of orange-red colored spermatozoa in the experiment group were increased significantly at 5 and 15 min respectively after uPA incubation (P < 0.05). (2) The mean fluorescence intensity of JC-1 within the spermatozoal body at 15, 30 and 60 min and the percentage of orange-red colored spermatozoa at 5 and 15 min in the group were significantly higher than those in the control group (P < 0.05). CONCLUSION: uPA could increase the mitochondrial membrane potential of mouse capacitated-spermatozoa in vitro, and maintain it at a high level for a certain period of time. By enhancing sperm mitochondrial function, uPA may provide sufficient energy for capacitated-spermatozoa to increase their motility and change their motor pattern, which might be one of the therapeutic mechanisms of uPA on male infertility.

The expression and significance of CATSPER1 in human testis and ejaculated spermatozoa.

AIM: To investigate the distribution of cation channel of sperm 1 (CATSPER1) protein and the presence of CATSPER1 mRNA in human testis and ejaculated spermatozoa. The influence of anti-human CATSPER1 antibody upon human sperm motility was used to evaluate the function of human CATSPER1 and to estimate its possible use as a target for immunocontraception. METHODS: Human ejaculated sperm from normozoospermic donors (n = 12) and liquid nitrogen frozen human testis were used for the study of mRNA and protein expression of CATSPER1 by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry, respectively. Spermatozoa from normozoospermic donors (n = 12) were individually processed using a swim-up procedure and were then incubated with CATSPER1 antibody at final concentrations of 20, 4 and 0.8 microg/mL. After 1, 2 and 6 h incubation, progressive motility and fast progressive motility were measured by means of computer-assisted semen analysis. RESULTS: CATSPER1 transcript was detected in both human testis and each human ejaculated semen sample. CATSPER1 protein expressed in the membrane of spermatid and was localized in the principal piece of the sperm tail. The application of CATSPER1 antibody at all concentrations significantly inhibited both progressive motility and fast progressive motility after 1, 2 and 6 h incubation, and significant dose-dependent changes were observed. CONCLUSION: CATSPER1 is meiotically and post-meiotically expressed in human testis tissue. CATSPER1 mRNA in human ejaculated spermatozoa could be a more feasible target for study and infertility screening than testis biopsy. In addition, our results suggest that human CATSPER1 could be a possible target for immunocontraception.

[Effects of testis murine cytomegalovirus infection on sperm acrosome reaction and spermatic function of membrane in mice].

OBJECTIVE: To explore the effects of testis murine cytomegalovirus (MCMV) infection on mature sperm acrosome reaction and spermatic function of membrane in mice. METHODS: BALB/c mice without MCMV infection were randomly devided into two groups: an experimental group (48 mice) and a control group (30 mice). The mice in the control group were treated by inoculation DMEM without MCMV into testis, while those in the experimental one were directly inoculated with MCMV into testis. Mice in two groups were sacrificed separately at 1, 2, 4, 6, 9, 14 d post inoculation (D1, 2, 4, 6, 9, 14 PI), and the MCMV M83 mRNA gene was detected inside the testes by in situ hybridization (ISH) with one episode MCMV late-mRNA probe labeled with digoxin, meanwhile acrosome reaction and the function of membrane of mature sperms in the epididymis tails was measured. RESULTS: The positive signal of ISH of MCMV was mainly founded in the two kinds of testicular cells (spermatogenic cells and Leydig cells) in the experimental group. Compared with that of the control group, the sperm acrosome reaction in the experimental group was decreased significantly by the rate from (71 +/- 6)%, (70 +/- 7)% to (58 +/- 9)%, (56 +/- 9)% (P < 0.05) separately on D2 PI and D4 PI. And the sperm membrane hypo-osmotic swelling was decreased significantly by the rate from (60 +/- 7)%, (50 +/- 4)% to (48 +/- 9)%, (38 +/- 8)% (P < 0.05) separately also on D2 PI and D4 PI. CONCLUSION: The model of CMV infection in murine testis was established. The sperm acrosome reaction and function of membrane in mice might be descented significantly by MCMV infection in the early period, which shows that MCMV infection might influent the sperm's function.