Upregulation of NFKBIZ affects bladder cancer progression via the PTEN/PI3K/Akt signaling pathway.

NF­κB inhibitor ζ (NFKBIZ), a member of the IκB family that interacts with NF­κB, has been reported to be an important regulator of inflammation, cell proliferation and survival. However, the role of NFKBIZ in bladder cancer (BC) remains unknown. The present study aimed to investigate the functions of NFKBIZ in BC. First, the expression levels of NFKBIZ and the associations between NFKBIZ expression and the clinical survival of patients were determined using BC tissue samples, BC cell lines and datasets from different databases. Two BC cell lines (T24 and 5637) were selected to overexpress NFKBIZ, and the proliferative, migratory and invasive abilities of cells were determined; additionally, tumor growth following transplantation in in vivo mouse models was analyzed using T24 cells overexpressing NFKBIZ. Subsequently, the association between NFKBIZ and PTEN was determined using data from databases and immunohistochemistry analysis of clinical and nude mice tumor tissues. Finally, the interactions between NFKBIZ, PTEN and the downstream PI3K/AKT/mTOR signaling pathway were evaluated using western blotting. In conclusion, the present results indicated that NFKBIZ expression was low in BC, and NFKBIZ inhibited the proliferation of BC cells through the PTEN/PI3K/Akt signaling pathway, suggesting that NFKBIZ may represent a novel prognostic biomarker in BC and may provide a potential therapeutic tumor­associated antigen for BC.

[Effect of testicular autophagy on spermatogenic cells in varicocele rats].

OBJECTIVE: To study the effect of different levels of autophagy in the testis on the apoptosis of spermatogenic cells in the rat model of varicocele (VC). METHODS: We randomly divided 54 SD male rats into six groups, blank control (n = 6), rapamycin control (n = 6), chloroquine control (n = 6), VC model control (n = 12), VC + rapamycin (n = 12), and VC + chloroquine (n = 12). We observed the histomorphological changes of the testis and epididymis by HE staining, obtained the scores on spermatogenesis in the testis and epididymis, calculated the apoptosis index (AI) of the testicular spermatogenic cells by TUNEL, and determined the expressions of LC3-Ⅱ, LC3-Ⅰ, p62, Bax and Bcl-2 proteins in the testis tissue by Western blot. RESULTS: There were no significant morphological changes in the testis and epididymis of the rats in the blank control, rapamycin control and chloroquine control groups, or significant differences in the scores on testicular and epididymal spermatogenesis and AI of the testicular spermatogenic cells (P>0.05). The animals in the VC model control group exhibited significant pathological damage in the testicular and epididymal tissues, with remarkably decreased scores on spermatogenesis (P<0.01) and increased AI (P<0.01), which were markedly improved in the VC + rapamycin group and slightly aggravated in the VC + chloroquine group compared with the VC model controls. In comparison with the rats in the blank control group, those in the VC model control group showed significantly up-regulated expressions of the autophagy-related protein LC3 (including the LC3-Ⅱ/LC3-Ⅰ ratio) and the pro-apoptotic protein Bax in testicular tissue (P<0.01) but down-regulated expression of the anti-apoptotic protein Bcl-2 (P<0.01). The expressions of LC3 and Bcl-2 in the testis tissue were significantly higher in the VC + rapamycin (P<0.01) but lower in the VC + chloroquine group (P<0.01), while those of p62 and Bax remarkably lower in the VC + rapamycin (P<0.01) but higher in the VC + chloroquine group than in the VC model controls (P<0.01). CONCLUSIONS: Varicocele induces autophagy in the testis and apoptosis of spermatogenic cells in rats. Up-regulating autophagy can inhibit while blocking autophagy can promote the apoptosis of spermatogenic cells.

TLR4 promotes the expression of HIF-1α by triggering reactive oxygen species in cervical cancer cells in vitro-implications for therapeutic intervention.

The present study investigated the mechanism underlying Toll-like receptor 4 (TLR4)-mediated stimulation of hypoxia-inducible factor-1α (HIF-1α) activity and its association with reactive oxygen species (ROS) in cervical cancer cells. SiHa cells were cultured and randomized to control, lipopolysaccharide (LPS), methyl-ß-cyclodextrin (MßCD)+LPS, ammonium pyrrolidinedithiocarbamate (PDTC)+LPS, ST2825+LPS and small interfering (si) RNA TLR4+LPS treatment groups. Cell proliferation was quantified using an MTT assay, cell cloning was performed using soft agar colony formation and HIF-1α expression was detected by immunocytochemical staining and western blot analyses. Dichloro-dihydro-fluorescein diacetate and lucigenin luminescence assays were used to detect alterations in ROS and nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase content, respectively. Co-localization of TLR4 and HIF-1α was detected by immunofluorescence staining and observed using fluorescence microscopy. Compared with the control group, cell proliferation was enhanced in the LPS-treated group and was not altered in the PDTC+LPS treatment group. Cell proliferation was reduced in all other treatment groups (P<0.05). Compared with the LPS group, cell proliferation decreased in all other groups. Compared with the PDTC+LPS treatment group, cell proliferation significantly decreased when LPS was co-administered with ST2825, siTLR4 and MßCD (P<0.01). Treatment with MßCD+LPS exhibited an increased inhibitory effect on cell activity and proliferation. Compared with the control group, HIF-1α expression was enhanced following treatment with LPS, although it decreased when LPS was co-administered with ST2825, siTLR4 and MßCD (P<0.05). HIF-1α expression decreased following treatment with ST2825, siTLR4, MßCD and PDTC+LPS, compared with treatment with LPS alone. Compared with the PDTC+LPS group, HIF-1α activity decreased when LPS was co-administered with ST2825, siTLR4 and MßCD. NADPH oxidase and ROS levels increased in cells treated with LPS, compared with the control group, at 24 and 12 h following treatment, respectively, and decreased at 12 h when LPS was co-administered with ST2825, siTLR4 and MßCD. There was no difference between the LPS and PDTC+LPS groups with respect to NADPH and ROS levels. Compared with the PDTC+LPS group, NADPH oxidase activity and ROS content decreased when LPS was co-administered with ST2825, siTLR4 and MßCD. NADPH oxidase activity and ROS content were lowest in the MßCD+LPS treatment group, and immunofluorescent staining demonstrated that TLR4 was localized to the cell surface and HIF-1α was primarily localized to the cytoplasm. TLR4 was co-expressed with HIF-1α in cervical cancer cells. The results of the present study suggested that TLR4 signaling primarily promoted HIF-1α activity via activation of lipid rafts/NADPH oxidase redox signaling and may be associated with the initiation and progression of cervical cancer. This promoting effect was stronger in TLR4/lipid rafts/NADPH oxidase pathway than that in TLR4-NF-κB signaling pathway. Therefore, the TLR4/lipid raft-associated redox signal may be a target for therapeutic intervention to prevent the growth of cervical cancer.

Effect of varicocelectomy treatment on spermatogenesis and apoptosis via the induction of heat shock protein 70 in varicocele­induced rats.

In the current study, the hypothesis that testicular varicocelectomy improves spermatogenesis and attenuates apoptosis via the induction of heat shock protein 70 (Hsp70) in a rat model of varicocele was investigated. Adult male Wistar rats (n=75) were randomly divided into 5 groups of 15 each: Control, sham, varicocele, varicocelectomy, and varicocelectomy plus Quercetin. A total of 6 weeks after the varicocelectomy, the left testis of all rats was removed for subsequent examination. Histological changes were compared between the groups. The expression of Hsp70 and apoptosis­associated indicators were evaluated based on immunohistochemical, western blot and mRNA expression analyses. Compared with the varicocele group, the varicocelectomy group exhibited a markedly reduced Bcl­2­associated X protein/B­cell lymphoma 2 (Bax/Bcl­2) ratio, and had a decreased expression of caspase­9, cytochrome c (cyt c) and caspase­3 through the intrinsic signal transduction pathways. Quercetin treatment inhibited the protective effects of varicocelectomy. The expression of Hsp70 was increased in the varicocele group which was further elevated by the varicocelectomy. These results indicated that varicocelectomy can reduce the Bax/Bcl­2 ratio, and decrease the levels of caspase­9, cyt c and caspase­3 via the mitochondrial signal transduction pathway. Such protective effects on left testis spermatogenesis and against apoptosis may be due to the induction of Hsp70. The findings of the present study suggested that varicocelectomy has a clear advantage in protecting testicular function and ameliorating spermatogenic cells apoptosis.

Autophagy may play an important role in varicocele.

The present study aimed to determine the expression of autophagy and investigate whether the hypoxia­inducible factor 1α (HIF­1α)/BCL2 interacting protein (BNIP3)/Beclin­1 autophagy signaling pathway serves an important role in activating autophagy in varicocele (VC) rat testes cells. Furthermore, the current study aimed to explain the possible association between autophagy and apoptosis. A total of 48 adult male Sprague Dawley rats were divided into group A (control), group B (VC 15­day), group C (VC 30­day) and group D (VC 45­day), with 12 rats in each group. The rats in group A did not receive any interventions, and in groups B, C, and D the VC model was established simultaneously. At 0, 15, 30, and 45 days, an orchidectomy on the left testes was performed in groups A­D, each on its respective day. Transmission electron microscopy was used to investigate the expression of autophagy. Compared with groups A and B, it was demonstrated that the expression of autophagy in groups C, and D was significantly increased. Hematoxylin and eosin staining revealed that as the rats survived VC longer, the testicular tissue damage became more serious. Furthermore, the Johnson score revealed that VC impaired the spermeiogenesis function of the male rats. Additionally, it was demonstrated that the apoptosis index of the semini-ferous epithelia cells in VC rat testes increased over time, as measured using TUNEL staining. Immunohistochemical analysis revealed that as the VC was prolonged, the expression of HIF­1α gradually increased while the expression of (apoptosis regulator Bcl­2) Bcl­2 gradually decreased. Furthermore, western blot analysis revealed that the protein expression of Bcl­2 decreased and apoptosis regulator Bax increased. Furthermore, HIF­1α, BNIP3, Beclin1 and microtubule associated protein 1 light chain 3 α (LC3)II/LC3I expression gradually increased. However, significant increases in Beclin 1 and LC3II/LC3I were only observed between the day 0 and day 30 groups. In addition, the expression of p62 significantly increased between day 0 and day 15, but gradually decreased between day 15 and day 45. The results of the present study revealed that VC can lead to testicular tissue hypoxia, and that the HIF­1α/BNIP3/Beclin1 autophagy signaling pathway may upregulate autophagy in VC rats testes. Thus, the association between autophagy and apoptosis may serve an important role in male infertility caused by VC.

Correlation of the occurrence of YMDD mutations with HBV genotypes, HBV-DNA levels, and HBeAg status in Chinese patients with chronic hepatitis B during lamivudine treatment.

BACKGROUND: Continuous lamivudine therapy is associated with high rates of YMDD mutations, which are the main causes of drug resistance. The current study explores the association of the emergence of YMDD mutations with pretherapy HBV genotype, HBV-DNA levels, HBeAg status, and serum alanine aminotransferase (ALT) levels in Chinese patients receiving lamivudine therapy for chronic hepatitis B. METHODS: A total of 319 chronic hepatitis B patients who received lamivudine therapy for more than a year were enrolled in this study. YMDD mutations, HBV genotype, HBV-DNA levels, HBeAg status, and ALT levels were determined prior to their lamivudine treatment and every three months for a year of this therapy. RESULTS: Among the 319 patients, 137 (42.95%) were infected with genotype B and 182 (57.05%) with genotype C. Up to 94 patients (29.47%) developed YMDD mutations within one year of lamivudine therapy. Furthermore, 50 patients with HBV genotype B and 44 patients with genotype C developed YMDD mutations (36.50% vs 24.18%, P<0.05). Logistic regression analysis showed that pretherapy HBV genotype, HBV-DNA levels, and HBeAg status are independent factors for the emergence of YMDD mutations (HBV genotype: OR=2.159, 95% CI 1.291-3.609, P=0.003; HBV-DNA: OR=1.653, 95% CI 1.231-2.218, P=0.001; HBeAg: OR=2.021, 95% CI 1.201-3.399, P=0.008). CONCLUSIONS: HBV genotype, HBV-DNA levels, and HBeAg status at baseline are the independent factors associated with the emergence of YMDD mutations among Chinese patients receiving lamivudine therapy for chronic hepatitis B. These findings are helpful to the development of therapeutic strategies for these patients.

[Electrophysiologial evaluation of ozone in treating of lumbar disc herniation with curving sheath-needle multi-direction rotating injection].

OBJECTIVE: To compare electrophysiological changes in treating lumbar disc herniation (LDH) with ozone by curving sheath-needle multi-direction rotating injection (CSNMRI) and conventional injection method. METHODS: From May 2005 to June 2009,100 patients with LDH were studied, included 68 males and 32 females, ranging in age from 25 to 58 years with an average of 44 years, in course of disease from 3 months to 8 years with an average of 8.8 months. All patients were numbered according to sequence of visit, and were completely randomly divided into group A and group B with DPS software, 50 cases in each group. All patients were injected ozone into lesion of intervertebral disc, in group A with CSNMRI and in group B with conventional method. The electrophysiologic study of all patients was performed respectively before treatment and at the 3 month after treatment. The electromyogram (EMG) of the main muscle groups of involved lower limb and the corresponding segments of sacrospinal muscle was tested; the duration and multiphase-wave rate of MUP were calculated. H-reflex of tibial nerve in both lower limbs was observed and the number of abnormal H-reflex and the H-wave latency were recorded. RESULTS: After treatment, the number of muscles with abnormal EMG was reduced to different degrees in each group, but there was more significant reduction in group A (P < 0.05 or 0.01); the duration and multiphase-wave rate of MUP in the two groups were both reduced and close to the normal level (P < 0.01), yet the changes in group A was more than that of group B (P < 0.05 or 0.01). There was no significant difference in the number of abnormal H-reflex before treatment between two groups, whereas was markedly lower in group A than that of group B after treatment (P < 0.05). After treatment, H-wave latency in two groups was shortened and become close to normal, but group B was more statistically significant than group B (P < 0.05). CONCLUSION: The neural electrophysiological abnormalities can reflect the degree of nerve root compression and damage, and is one of the objective indicators to estimate neuromuscular function. It can better meliorate abnormal electrophysiology to inject ozone to treat LDH with CSNMRI than conventional method.

Determination of serum levels of YKL-40 and hyaluronic acid in patients with hepatic fibrosis due to schistosomiasis japonica and appraisal of their clinical value.

The objectives of this study were (i) to evaluate the serum levels of human cartilage glycoprotein 39 (YKL-40) and hyaluronic acid (HA) in people infected with Schistosoma japonicum, and (ii) to determine their clinical value. A total of 563 people were subjected to ultrasonography, and 60 patients were identified with either mild (n=30) or severe (n=30) hepatic fibrosis. In addition, 28 healthy subjects were included as controls. Blood sera of these 88 people were examined with regard to the levels of YKL-40 and HA. The former was measured by enzyme-linked immunosorbent assay, and serum HA was determined by a commercially available radioimmunoassay method. On the basis of the ultrasonographic investigations, HA levels in normal, mild, and severe cases of hepatic fibrosis were 83.0+/-35.7, 216.1+/-77.9 and 212.6+/-80.9 microg/ml, respectively. When the same sera were tested for YKL-40, 49.0+/-10.4, 92.3+/-18.5 and 172.1+/-35.9 microg/ml, respectively, were recorded in the three groups. Thus, the serum levels of YKL-40 are not only increased in patients infected with S. japonicum but they are also correlated with the stage of hepatic fibrosis. In conclusion, it appears that YKL-40 is more sensitive than HA in measuring the degree of hepatic fibrosis due to schistosomiasis, which warrants further investigation.

Positive association between synapsin II and schizophrenia.

BACKGROUND: Synapsin II encodes a neuron-specific phosphoprotein that selectively binds to small synaptic vesicles in the presynaptic nerve terminal. The expressions of messenger ribonucleic acid and protein of synapsin II have been reported to be significantly reduced in the brains of schizophrenia patients. The synapsin II gene is located on 3p25, a region that has been implicated to be associated with schizophrenia by genetic linkage. All these findings suggest synapsin II as a candidate gene for schizophrenia. METHODS: In this work, we studied four markers (two single nucleotide polymorphisms (SNPs): rs308963 and rs795009; and two insertion/deletion polymorphisms: rs2307981 and rs2308169) covering 144.2 kilobase pairs (kb) with an average interval of 38 kb in synapsin II in a sample of 654 schizophrenic patients and 628 normal control subjects to explore the mechanism underlying schizophrenia. RESULTS: We found significant differences in allele frequency distribution of SNP rs795009 (p =.000018, odds ratio 1.405, 95% confidence interval 1.202-1.641) between patients and control subjects. The T allele was significantly higher in patients than in control subjects. Moreover, the overall frequency of haplotype showed significant differences between patients and control subjects (p <.000001). CONCLUSIONS: This study suggests a positive association between synapsin II and schizophrenia, implying that synapsin II is involved in the etiology of schizophrenia.

[Study on the disabilities in aged 0-7 years children in Shenzhen, China].

OBJECTIVE: To explore the prevalence of vision, mental, audibility, language, psychiatry, extremity, and influence factors in the 0 - 7 year olds. METHODS: A total number of 77,727 0 - 7 year old children living in Shenzhen city were tested with tree phase screening under the Chinese standard of evaluation in disabilities. RESULTS: The prevalence of all disabilities was 5.59 per thousand (adjusted rate was 8.49 per thousand with a false negative of 3.1 per thousand ). The prevalence of mental disease was the highest (1.88 per thousand, with adjusted rate 3.43 per thousand ), the prevalence of language disability was 1.88 per thousand (including retarded language development, with adjusted rate 3.43 per thousand ). The prevalence rates of psychiatry, extremity and audibility disability were 1.59 per thousand, 1.56 per thousand, 1.11 per thousand respectively with of vision the lowest (0.37 per thousand ). The prevalence of all disabilities, audibility, language and mental was on the increase with age. The difference was statistically significant. Among all different age groups regarding psychiatric disease, the highest fell in the 2 - 4 year olds. The prevalence of extremity was not statistically different among age groups. The suspected agents of disease which occurred before or during pregnancy took up 45.7%. CONCLUSION: The prevalence of six kinds disabilities in Shenzhen was about 10 per thousand lower than that of the samples of the nation in 1989, but two times higher than that of similar studies in Japan. The prevalence rates of language and psychiatric disease were higher than that of the nation in 1989. The causation should be further studied.