Relationship between Erbin and Bax/Bcl-xL-mediated cell apoptosis during sepsis-induced acute kidney injury in mice / 中华麻醉学杂志

Objective:To evaluate the relationship between Erbin and Bax/Bcl-xL-mediated cell apoptosis during sepsis-induced acute kidney injury in mice.Methods:Thirty-two SPF male wild type C57BL/6 mice, 32 SPF male Erbin (-/-) C57BL/6 mice, aged 6-8 weeks, weighing 20-30 g, were divided into 2 groups ( n=16 each) using the random number table method: wild type sham operation group (WT+ Sham group), wild type sepsis group (WT+ S group), Erbin(-/-) sham operation group (EKO+ Sham group), and Erbin(-/-) sepsis group (EKO+ S group). The sepsis model was established using the moderate cecal ligation and puncture (CLP) in anesthetized animals.The survival rates within 7 days after CLP were recorded.The serum concentrations of tumor necrosis factor-alpha (TNF-α), interleukin-10 (IL-10), IL-1β, creatinine (Cr), blood urea nitrogen (BUN) and lactic dehydrogenase (LDH) were determined at 24 h after CLP.Then the renal tissues were taken for assessment of renal injury which was scored and for determination of the apoptosis rate (by TUNEL) and expression of cleaved-caspase-3, Bcl-xL and Bax (by Western blot). Results:Compared with sham operation groups, the survival rates were significantly decreased, the serum concentrations of IL-1β, IL-10, TNF-α, Cr, BUN and LDH, renal injury score and apoptosis rate were increased, the expression of Bax and cleaved-caspase-3 was up-regulated, and the expression of Bcl-xL was down-regulated in sepsis groups ( P<0.05). Compared with WT+ S group, the survival rates were significantly decreased, the serum concentrations of IL-1β, LDH, TNF-α, Cr and BUN and renal injury score were increased, the serum concentration of IL-10 was decreased, the apoptosis rate of renal tissues was increased, the expression of Bax and cleaved-caspase-3 was up-regulated, and the expression of Bcl-xL was down-regulated in EKO+ S group ( P<0.05). Conclusions:Erbin can inhibit Bax/Bcl-xL-mediated cell apoptosis and is involved in endogenous protective mechanism against sepsis-induced acute kidney injury in mice.

BCL-X expression in oral cancer: Comparison between oral squamous cell carcinoma and verrucous carcinoma

Verrucous carcinoma (VC) should be considered a distinct clinicopathologic entity different from the more common oral squamous cell carcinoma (OSCC) because of its unique biological behavior. Best way to understand the behavior of these carcinomas is to study them by means of molecular methods, especially in tumor progression tests and Bcl-X is an important antiapoptotic member of the Bcl-2 family and is one of the newest and most useful markers to determine the aggressiveness of many carcinomas. The relationship between this Bcl-X protein and carcinomatous behavior toward it is not studied extensively, which we attempted to evaluate using immunohistochemical analysis in selected carcinomas of the head and neck region. Method: We studied Bcl-X protein expression in sections of thirty OSCC and ten VC samples and correlated this with tumor differentiation. Results: There was a significant difference in cytoplasmic staining of Bcl-X expression with statistical analysis (P < 0.005) for VC and OSCC when compared as a group. No significance was seen among the different histological grades of OSCC and when compared with VC individually. Conclusion: The significant result between OSCC and VC suggests that their biologic course is comparable and can be helpful in differentiating them with each other for establishment of a better treatment protocol.

Keratinization of Lung Squamous Cell Carcinoma Is Associated with Poor Clinical Outcome / 결핵및호흡기질환

BACKGROUND: Although the World Health Organization (WHO) classification of lung squamous cell carcinoma (SCC) was revised in 2015, its clinical implications for lung SCC subsets remain unclear. We investigated whether the morphologic characteristics of lung SCC, including keratinization, were associated with clinical parameters and clinical outcome of patients. METHODS: A total of 81 patients who underwent curative surgical resection of diagnosed lung SCC, were enrolled in this study. Attributes such as keratinization, tumor budding, single cell invasion, and nuclear size within the tumor, as well as immunohistochemistry of Bcl-xL and pS6 expressions, were evaluated. RESULTS: The keratinizing and nonkeratinizing subtypes did not differ with respect to age, sex, TNM stage, and morphologic parameters such as nuclear diameter, tumor budding, and single cell invasion at the tumor edge. Most patients with the keratinizing subtype (98.0%) had a history of smoking, whereas the nonkeratinizing group had a relatively higher proportion of never-smokers relative to the keratinizing group (24.0% vs. 2.0%; p=0.008, chi-square test). Expression of pS6 (a surrogate marker of mammalian target of rapamycin complex 1 [mTORC1] signaling that regulates keratinocyte differentiation), and Bcl-xL (a key anti-apoptotic molecule that may inhibit keratinization), did not correlate significantly with the presence of keratinization. Patients with the keratinizing subtype had a significantly shorter overall survival (85.2 months vs. 135.7 months, p=0.010, log-rank test), and a multivariate analysis showed that keratinization was an independent, poor prognostic factor (hazard ratio, 2.389; 95% confidence interval, 1.090–5.233; p=0.030). CONCLUSION: In lung SCC, keratinization is associated with a poor prognosis, and might be associated with smoking.

INXS, um longo RNA não codificador de proteínas mediador da apoptose / INXS, a long noncoding RNA that mediates apoptosis

O splicing alternativo do pré-mRNA de BCL-X produz duas isoformas de mRNAs com funções antagônicas, a pró-apoptótica BCL-XS e a anti-apoptótica BCL-XL, cujo balanço regula a homeostasia celular. Entretanto, o mecanismo que regula esse processamento ainda é desconhecido. Nesse trabalho, nós identificamos e caracterizamos um longo RNA não codificador de proteínas (lncRNA) nomeado INXS, que é transcrito a partir da fita oposta do locus genômico de BCL-X, sendo menos abundante em linhagens celulares tumorais e tecidos tumorais de pacientes quando comparados com os respectivos pares não tumorais. INXS é um RNA unspliced de 1903 nts, é transcrito pela RNA Polimerase II, possui cap 5', está enriquecido na fração nuclear das células e se liga à proteína Sam68 do complexo modulador de splicing. O tratamento de células tumorais 786-O com cada um de três agentes indutores de apoptose aumentou a expressão endógena do INXS, levando ao aumento expressivo da proporção entre os mRNAs de BCL-XS / BCL-XL, e ativação das caspases 3, 7 e 9. Estes efeitos foram anulados na presença do knockdown do INXS. Da mesma forma, a superexpressão ectópica do INXS causou uma mudança no splicing favorecendo a isoforma BCL-XS e ativação das caspases, aumentando os níveis da proteína BCL-XS e conduzindo as células à apoptose. Utilizando um modelo in vivo, cinco injeções intra-tumorais do INXS durante 15 dias causaram uma regressão acentuada no volume dos xenotumores. Portanto, INXS é um lncRNA que induz a apoptose, sugerindo que essa molécula seja um possível alvo a ser explorado na terapia contra o câncer

BCL-X mRNA alternative splicing generates pro-apoptotic BCL-XS or anti-apoptotic BCL-XL, whose balance regulates cell homeostasis. However, the mechanism that regulates the splice shifting is incompletely understood. Here, we identified and characterized a long noncoding RNA (lncRNA) named INXS, transcribed from the opposite genomic strand of BCL-X, that was less abundant in tumor cell lines and patient tumor tissues compared with non-tumors. INXS is an unspliced 1903 nt-long RNA, is transcribed by RNA Polymerase II, 5'-capped, nuclear enriched and binds Sam68 splicing-modulator. The treatment of tumor cell line 786-O with each of three apoptosis-inducing agents increased endogenous INXS lncRNA, increased BCL-XS / BCL-XL mRNA ratio, and activated caspases 3, 7 and 9. These effects were abrogated in the presence of INXS knockdown. Similarly, ectopic INXS overexpression caused a shift in splicing towards BCL-XS and activation of caspases, increasing the levels of BCL-XS protein and then leading the cells to apoptosis. In a mouse xenograft model, five intra-tumor injections of INXS along 15 days caused a marked regression in tumor volume. INXS is an lncRNA that induces apoptosis, suggesting that INXS is a possible target to be explored in cancer therapies

Study on the inhibitory effect of the black garlic extract on the growth of hela cells / 中国医师杂志

Objective To explore the effect of black garlic extract on HeLa cells and its possible molecular mechanisms.Methods Inhibitory rate of HeLa cells was detected with methyl thiazolyl tetrazolium(MTT).Flow cytometry annexin V-fluorescein isothiocyanate/propidium iodide (V-FITC/PI) was used to detect tumor cell apoptosis.Flow cytometry was used to measure cell-cycle changes.Immunohistochemistry method was used to detect expressions of tumor proteins bax and Bcl-2.Results Black garlic extract significantly inhibited the growth of HeLa cells.Black garlic extract significantly induced apoptosis of HeLa cells.The apoptosis rate was (53.26 ± 1.78)％ in the black garlic high-dose group,and (3.68 ±0.11)％ in the control group.Black garlic extract affected cell cycle,upregulated bax expression,and downregulated bcl-2 expression.Conclusions Black garlic extract significantly induced the apoptosis of HeLa cells.This effect might be caused through increasing G2/M cells or changing expressions of bax and bcl-2.

Effect of pioglitazone on the expression of PPARγ, NF-κB c-Rel and Bcl-xL in cultured rat cortical neurons after the oxygen-glucose/reoxygenation / 国际脑血管病杂志

Objective To investigate the effect of pioglitazone (PIO) on the expression of peroxisome proliferators-activated receptor γ (PPARγ),nuclear factor-κB (NF-κB) c-Rel and anti-apoptosis factor Bcl-xL in cultured Wistar rat cortical neurons after oxygen-glucose deprivation/reoxygenation (OGD/R).Methods The rat cerebral cortical neurons were primarily cultured in vitro and a model of OGD/R was induced.The rats were divided into 7 groups:normal,OGD 4 h/R 6 h,OGD 4 h/R 12 h,OGD 4 h/R 6 h + PIO,OGD 4 h/R 6 h + PPARγ inhibitor T0070907 (TIO),OGD 4 h/R 12 h + PIO,and OGD 4 h/R 12 h + TIO groups.Western blot was used to detect the expression of PPARγ and NF-κB c-Rel protein in neurons.Reverse transcriptionpolymerase chain reaction assay was used to detect the expression of Bcl-xL mRNA in neurons.Results Western blot analysis showed that the expression levels of PPARγ and NF-κB c-Rel proteins after OGD/R were increased significantly compared to those of the normal group (P ＜0.01).After giving PIO,the expression levels of PPARγ and NF-κB c-Rel proteins were further increased,and they were significantly higher than those in the OGD 4 h/R 6 h and OGD 4 h/R 12 h groups (P ＜0.01),and after giving TIO,the expression levels of PPARγ and NF-κB c-Rel proteins were decreased,and they were significantly lower than those in the OGD/R group (P ＜0.05) and the corresponding PIO group (P ＜0.01).Reverse transcription-polymerase chain reaction analysis showed that the expression level of Bcl-xL mRNA in the OGD 4 h/R 6 h group was significantly higher than that in the normal group (P ＜0.01).The expression level of Bcl-xL mRNA in the PIO group was significantly higher than that in the OGD 4 h/R 6 h and OGD 4 h/R 12 h groups (P ＜0.01).After giving TIO,the expression of Bcl-xL mRNA was decreased.It was significantly lower than that in the OGD/R and corresponding PIO groups (P ＜ 0.05 and P ＜ 0.01).Conclusions PIO can upregulate the expression of PPARγ protein in cultured cortical neurons after OGD/R,and then increase the expression of anti-apoptotic factor Bcl-xL mRNA of NF-κB c-Rel regulation.This may be the part mechanism of PPARγ neuroprotective effect.

Proteínas pro-apoptóticas y anti-apoptóticas como factores de pronóstico en Linfoma B difuso de célula grande en adultos / Pro-apoptotic and anti-apoptotic proteins as prognostic factors in diffuse large B-cell lymphoma in adults / Proteínas pró-apoptóticas e anti-apoptóticas como fatores de prognóstico no Linfoma B difuso de célula grande em adultos

Objetivo. El propósito de este estudio fue evaluar en pacientes con LBDCG la expresión de las proteínas anti-apoptóticas Bcl-2 y Bcl-X L y de las proteínas pro-apoptóticas Bad y Bax y su asociación con la supervivencia. Materiales y métodos. Se analizaron biopsias de 28 pacientes con diagnóstico de LBDCG. La expresión de los reguladores apoptóticos se evaluó mediante western blot. La asociación entre la expresión de las proteínas y la supervivencia fue analizada mediante el método de Kaplan-Meier y la prueba log-rank. Resultados. Las proteínas Bcl-2, Bak, Bad y Bcl-xL se encontraron expresadas en el 78,8%; 71,4%; 64,3% y 50% de los casos de LBDCG respectivamente. No encontramos asociación entre la presencia de las proteínas o sus niveles de expresión y la supervivencia total. La presencia de las proteínas Bad y Bcl-xL se asoció con una mayor supervivencia libre de enfermedad (33,3% vs. 20,0%, p LR test= 0,003; 42,9% vs. 14,3%, p LR test= 0,03 respectivamente). Niveles altos de expresión de Bad y de Bcl-X L se asociaron con una supervivencia libre de enfermedad mayor (35,7% vs. 21,4%, p LR test= 0,012 y 42,9% vs. 14,3%, p LR test= 0,045 respectivamente). Conclusión. Dado que la expresión de la proteína Bad en los tumores se asoció con una mayor supervivencia libre de enfermedad, los pacientes con bajos niveles de expresión de esta proteína podrían ser beneficiados en un futuro con terapias orientadas a inhibir las moléculas anti apoptóticas Bcl-xL y Bcl-2 mediante el empleo de moléculas que se unen específicamente al dominio BH3.

Objective. Our purpose was to evaluate the expression of antiapoptotic proteins Bcl-2 and Bcl-xL, and pro-apoptotic proteins Bad and Bax and their association with survival, in patients with DLBCL. Materials and methods. We analyzed biopsies from 28 patients diagnosed with DLBCL. The expression of the apoptotic regulators was assessed by western blot. The association between protein expression and survival was analyzed by the Kaplan-Meier method and the log-rank test. Results. Bcl-2, Bak, Bad and Bcl-xL proteins were expressed in 78.8, 71.4, 64.3 and 50% of the DLBCL cases, respectively. We found no association between the presence of proteins or their expression levels and overall survival. Both Bad and Bcl-xL were associated with higher disease-free survival (33.3% vs. 20.0%, p LR test= 0,003; 42.9% vs. 14.3%, p LR test= 0.03, respectively). High expression levels of Bad and Bcl-xL were associated with a higher disease-free survival (35.7% vs. 21.4%, p LR test= 0.012 y 42.9% vs. 14.3%, p LR test= 0.045, respectively). Conclusion. Given that expression of the Bad protein in tumors was related to a higher disease-free survival, patients with low expression levels of Bad could be candidates in future therapies oriented towards the inhibition of the anti-apoptotic proteins Bcl-xL and Bcl-2 by using molecules that bind specifically to the BH3 domain.

Objetivo. O objetivo deste estudo foi avaliar em pacientes com LBDCG a expressão das proteínas anti-apoptótica Bcl-2 e Bcl-X L e das proteínas pró-apoptóticas Bad e Bax e sua associação com a sobrevivência. Materiais e métodos. Foram analisadas biópsias de 28 pacientes com diagnóstico de LBDCG. A expressão de reguladores de apoptose foi avaliada por Western blot. A associação entre a expressão das proteínas e a sobrevivência foi analisada pelo método de Kaplan-Meier e o teste log-rank. Resultados. As proteína Bcl-2, Bak, Bad e Bcl-xL foram expressas em 78,8%, 71,4%, 64,3% e 50% dos casos de LBDCG, respectivamente. Nós não encontramos associação entre a presença das proteínas ou de seus níveis de expressão e a sobrevivência total. A presença das proteínas Bad e Bcl-xL foi associada à maior sobrevivência livre da doença (33,3% vs. 20,0%, p LR teste= 0,003; 42,9% vs. 14,3%, p LR teste= 0,03, respectivamente). Altos níveis de expressão de Bad e de Bcl-X L foram associados com uma sobrevivência livre da doença maior (35,7% vs. 21,4%, p LR teste= 0,012 e 42,9% vs. 14,3%, p LR teste= 0,045, respectivamente). Conclusão. Uma vez que a expressão da proteína Bad em tumores foi associada com uma maior sobrevivência livre de doença, os pacientes com baixos níveis de expressão dessa proteína poderiam ser beneficiados num futuro com terapias orientadas a inibir as moléculas anti-apoptóticas Bcl-xL e Bcl-2 utilizando moléculas que se ligam especificamente ao domínio BH3.

The expression and meaning of bcl-xL in superficial bladder tumor / 中国医师进修杂志

Objective To investigate the relations of the expression of bcl-xL in superficial bladder tumor and prognosis. Method The expression of bcl-xL was detected by envision system in 80 cases of superficial bladder tumor. Results In patients with high expression of bcl-xL, the recurrence rate was higher than that of normal expression [71.4%(30/42) vs 50.0%(19/38) ,P < 0.05], and the recurrence of time as early as normal expression [(16.0 ± 1.2) months vs (36.0 ± 4.5) months](P < 0.05). Conclusion The expression of bcl-xL could effectively predict the prognosis of patients with bladder tumor, those who with high expression of-bcl-xL have bad prognosis.

Calcium overload is essential for the acceleration of staurosporine-induced cell death following neuronal differentiation in PC12 cells

Differentiation of neuronal cells has been shown to accelerate stress-induced cell death, but the underlying mechanisms are not completely understood. Here, we find that early and sustained increase in cytosolic ([Ca2+]c) and mitochondrial Ca2+ levels ([Ca2+]m) is essential for the increased sensitivity to staurosporine-induced cell death following neuronal differentiation in PC12 cells. Consistently, pretreatment of differentiated PC12 cells with the intracellular Ca2+-chelator EGTA-AM diminished staurosporine-induced PARP cleavage and cell death. Furthermore, Ca2+ overload and enhanced vulnerability to staurosporine in differentiated cells were prevented by Bcl-XL overexpression. Our data reveal a new regulatory role for differentiation-dependent alteration of Ca2+ signaling in cell death in response to staurosporine.

Relationship and significance of HIF-1α and bcl-XL in gastric cancer / 肿瘤研究与临床

Objective To study the expression of HIF-1α and bcl-XL in gastric cancer and their relationship with tumor angiogenesis,clinicopathologic feature and prognosis.Methods Immunohistochemical technique was used to detect the expression of HIF-1α and bcl-XL in 54 cases of gastric canoer.SPSS 12.0 software was used to analyze the relationship between the expression of HIF-1α, bcl-XL and tumor angiogenesis,clinicopathologic feature of patients.Results The positive expression rate of HIF-1α in gastric cancer(74.07%) was significantly higher than that in normal gastric tissue(0),P＜0.01;The expression of HIF-1α in gastric cancer was significantly associated with TNM stage,invasive depth and lymph-node metastasis,P＜0.05 or P＜0.01;The positive expression rate of bcl-XL in gastric cancer(53.7%) was significantly higher than that in normal gastric tissue(33.3%),P＜0.05;The expression of bcl-XL in gastric cancer was significantly associated with histological types,TNM stage and lymph-node metastasis,P＜0.05;There was a positive correlation between expression of HIF-1α and bcl-XL (r=0.41,P＜0.05).Conclusion HIF-1α and bcl-XL play a very important role in the development in gastric cancer and could be a factor in diagnosis of gastric cancer and estimation of prognosis.

Inhibition of gamma ray-induced apoptosis by stimulatory heterotrimeric GTP binding protein involves Bcl-xL down-regulation in SH-SY5Y human neuroblastoma cells

Heterotrimeric GTP-binding proteins (G proteins) transduce extracellular signals into intracellular signals by activating effector molecules including adenylate cyclases that catalyze cAMP formation, and thus regulate various cellular responses such as metabolism, proliferation, and apoptosis. cAMP signaling pathways have been reported to protect cells from ionizing radiation-induced apoptosis, but however, the protective mechanism is not clear. Therefore, this study aimed to investigate the signaling molecules and the mechanism mediating the anti-apoptotic action of cAMP signaling system in radiation-induced apoptosis. Stable expression of a constitutively active mutant of G alpha s (G alpha sQL) protected gamma ray-induced apoptosis which was assessed by analysis of the cleavages of PARP, caspase-9, and caspase-3 and cytochrome C release in SH-SY5Y human neuroblastoma cells. G alpha sQL repressed the gamma ray-induced down-regulation of Bcl-xL protein, but transfection of Bcl-xL siRNA increased the gamma ray-induced apoptosis and abolished the anti-apoptotic effect of G alpha sQL. G alpha sQL decreased the degradation rate of Bcl-xL protein, and it also restrained the decrease in Bcl-xL mRNA by increasing the stability following ionizing irradiation. Furthermore, prostaglandin E2 that activates G alpha s was found to protect gamma ray-induced apoptosis, and the protective effect was abolished by treatment with prostanoid receptor antagonist specific to EP2/4R subtype. Moreover, specific agonists for adenosine A1 receptor that inhibits cAMP signaling pathway augmented gamma ray-induced apoptosis. From this study, it is concluded that Galphas-cAMP signaling system can protect SH-SY5Y cells from gamma ray-induced apoptosis partly by restraining down-regulation of Bcl-xL expression, suggesting that radiation-induced apoptosis can be modulated by GPCR ligands to improve the efficiency of radiation therapy.

Modification of Alternative Splicing of Bcl-x Pre-mRNA in Bladder Cancer Cells / 华中科技大学学报（医学）（英德文版）

To modify the splicing pattern of Bcl-x and compare the effect of this approach with that of the antisense gene therapy in BIU-87 cell line of bladder cancer, by using 5'-Bcl-x AS to target downstream alternative 5'-Bcl-x splice site to shift splicing from Bcl-xL to Bcl-xS and 3'-Bcl-x AS antisense to the 3'-splice site of exon Ⅲ in Bcl-x pre- mRNA to down regulation of Bcl-xL expression,the inhibitory effects on cancer cells by modification of alternative splicing and antisense gene therapy were observed and compared by microscopy, MTT Assay, RT-PCR, FACS, Westhern bloting and clone formation. The growth of cells BIU-87 was inhibited in a dose- and time-dependent manner. Its inhibitory effect began 12 h after the exposure, reaching a maximum value after 72h. The number of cells decreased in S phase and the number increased in G1 phase. The ability to form foci was reduced and the antisense gene therapy was approximately half as efficient as modification of alternative splicing in inducing apoptosis. It is concluded that modification of splicing pattern of Bcl-x pre-mRNA in bladder cancer cell BIU-87 is better than antisense gene therapy in terms of tumor inhibition.

Effect of 60% oxygen exposure on expression of Bax and Bcl-X_L in lungs of newborn rats / 中华围产医学杂志

Objective To investigate the effect of Bax and Bcl-X_L expression in newborn rat with moderate hyperoxic exposure. Methods Hyperoxic lung injury model was established by exposure to 60% O_2 in the neonatal period of SD rats. Rats exposed to air were used as control groups, with 8 animals in each group on repeated experiments. The pathology of pulmonary tissues was detected by HE stain. Mean alveolar area and alveolar number per ?m~2 were applied to estimate the pathological effects of prolonged hyperoxia in neonatal rats. The expression of Bax and Bcl-X_L proteins in lung were detected by immunohistochemistry and the expressions of Bax and Bcl-X_L mRNA by RT-PCR. Results In hyperoxia groups, alveolar dysplasia appeared 4 days after hyperoxia, mean alveolar area increased and alveolar number per ?m~2 decreased from the 4th day. Bax and Bcl-X_L protein were mainly expressed on bronchiolar epithelial cells and vascular endothelial cells. Compared with control group, the expression of Bax increased from the 1st day after hyperoxia, Bax mRNA decreased from the 11th day (q=8.4802, P

Effects of Endothelin A Receptor Antagonist to Cellular Proliferation and Apoptosis in Neonatal Rat Heart

PURPOSE: In order to investigate the role of endothelins in the cardiac development, the present study was designed to examine the effects of endothelin A receptor(ETAR) antagonist to the cellular proliferation and apoptosis in the neonatal rat heart. In addition, the expression of various regulatory genes in protein and mRNA levels by ETAR antagonist were examined. METHODS: Neonatal Spargue-Dawley rats were separated into two groups. The BMS group(N=22) was treated with the selective ETAR antagonist(Bristo-Myers Squibb-182874; 300 mg/Kg/day) and the control group(N=20) with normal saline for seven days by orogastric tube. On the following day, their hearts were harvested for determination of apoptosis by modified TUNEL technique and cellular proliferation by PCNA stain. In addition to this, Western blottings and RT-PCRs of bcl-x, clusterin, p53 and TGF-beta1 were performed. RESULTS: The BMS group resulted in a reduced body weight, but not a significantly reduced heart weight. In the BMS group, cardiac apoptotic cells and PCNA positive cells were decreased(P<0.05). In the BMS group, clusterin and bcl-x protein expressions were increased(P<0.05), but p53 and TGF-beta1 protein expressions remained the same. In the BMS group, clusterin and TGF-beta1 mRNA expressions were increased(P<0.05), but bcl-x and p53 mRNA expressions remained the same. CONCLUSION: ETAR antagonist treatment decreases cell turnover in the developing rat heart, which may account for the role of endothelins on modulating cardiac growth. These changes may be affected by clusterin and bcl-x expressions. These results support that there are some roles of endothelin and ETAR in the cellular level of early neonatal cardiac growth.

Apoptosis in the Human Neonatal Vascular Remodeling

BACKGROUND AND OBJECTIVES: In order to verify that apoptosis is one of the possible mechanisms of human neonatal vascular remodeling during transition from fetal to neonatal circulation, we identified apoptosis and analyzed its mechanism by evaluating apoptosis-related genes in umbilical vessel versus ascending aorta, ductus arteriosus (DA) versus adjacent pulmonary artery and aorta, and aorta versus its branching arteries. MATERIALS AND METHODS: Twenty-two umbilical cords, six ductus arteriosus with adjacent aortae and pulmonary arteries, and four aortic arches with their branching great arteries, were obtained from neonates. The presence of apoptotic cells was demonstrated by electron microscopy (EM) and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). Immunohistochemical staining and Western blotting were used for the analysis of the proteins of apoptosis-related gene. RESULTS: Apoptosis of the smooth muscle cells of the umbilical vessels were identified in all UC, which were examined by electron microscopy and TUNEL. The expressions of Bax and Bcl-X were shown by immunohistochemistry to be stronger in umbilical artery than in the neonatal aorta, but Bcl-2 was weak in both arteries. In the immunoblot analysis of UC, the expression of the proapoptotic short isoform of Bcl-X was stronger than in other tissue, and caspase-3 was selectively activated whereas it was not in the other components of the cardiovascular system. However, the expression patterns of the Fas Ag and Fas ligand (i.e. positive Fas Ag and negative Fas ligand), were similar in umbilical artery and aorta. This Bax-associated apoptosis was also observed in other vascular sites which undergo dramatic hemodynamic changes during birth, such as, the ductus arteriosus and the branching points of the great arteries from the aortic arch. CONCLUSIONS: Apoptosis is involved in the closure and regression of human umbilical vessels and the ductus arteriosus and in the remodeling of the branching great arteries during the neonatal period, where the Bax/Bcl-2/Bcl-X system, not Fas Ag/Fas ligand system, is likely to play a key role.

Up-regulation of Intragraft Gene Expression of Bcl-xL in Response to Injuries to Renal Allograft / 대한신장학회잡지

BACKGROUND: Bcl-xL and bax act as checkpoints of the apoptotic cell death. Although apoptosis is one of major mechanism of cell death in renal allografts inflicted by various events, the role of bcl-xL and bax in kidney transplantation has not been characterized yet. We therefore studied intragraft expression of bcl-x and bax and its clinical significance in renal transplantation. METHODS: We localized the expression of bcl-x, bcl-xS and bax proteins by immunohistochemistry, and measured the magnitude of the gene transcription of bax and bcl-xL by semi-quantitative RT-PCR in 37 implantation allograft biopsies(18 from 9 cadaveric donors, 19 from living donors), and 17 biopsies from patients undergoing acute rejection(AR). RESULTS: Immunoreactivities for bax, bcl-x, and bcl-xS were observed in tubular epithelial cells but not in glomeruli and vessels in implantation and AR biopsies. The infiltrating lymphocytes in AR expressed bax and bcl-xS but not for bcl-x. Comparing the intragraft gene transcript level of each allograft of a pair of recipients, who received graft from the same cadaveric donor, showed a higher bcl-xL in the patients with a higher concentration of postoperative 7th day serum creatinine. The transcript level of bcl-xL was higher in the Banff grade II and III AR biopsies than in the borderline or grade I AR, and also higher in steroid-resistant AR than in steroid-responsive patients. CONCLUSION: These results implicated the apoptotic death of infiltrating lymphocytes during rejection, and the compensatory up-regulation of bcl-xL in response to various apoptotic stimuli occurring in renal allografts.

Up-regulation of Intragraft Gene Expression of Bcl-xL in Response to Injuries to Renal Allograft / 대한신장학회잡지

BACKGROUND: Bcl-xL and bax act as checkpoints of the apoptotic cell death. Although apoptosis is one of major mechanism of cell death in renal allografts inflicted by various events, the role of bcl-xL and bax in kidney transplantation has not been characterized yet. We therefore studied intragraft expression of bcl-x and bax and its clinical significance in renal transplantation. METHODS: We localized the expression of bcl-x, bcl-xS and bax proteins by immunohistochemistry, and measured the magnitude of the gene transcription of bax and bcl-xL by semi-quantitative RT-PCR in 37 implantation allograft biopsies(18 from 9 cadaveric donors, 19 from living donors), and 17 biopsies from patients undergoing acute rejection(AR). RESULTS: Immunoreactivities for bax, bcl-x, and bcl-xS were observed in tubular epithelial cells but not in glomeruli and vessels in implantation and AR biopsies. The infiltrating lymphocytes in AR expressed bax and bcl-xS but not for bcl-x. Comparing the intragraft gene transcript level of each allograft of a pair of recipients, who received graft from the same cadaveric donor, showed a higher bcl-xL in the patients with a higher concentration of postoperative 7th day serum creatinine. The transcript level of bcl-xL was higher in the Banff grade II and III AR biopsies than in the borderline or grade I AR, and also higher in steroid-resistant AR than in steroid-responsive patients. CONCLUSION: These results implicated the apoptotic death of infiltrating lymphocytes during rejection, and the compensatory up-regulation of bcl-xL in response to various apoptotic stimuli occurring in renal allografts.

Preparation of gfp-bcl-X_L-contained recombinant adenovirus vector by the homologous recombination in bacteria / 中国病理生理杂志

AIM: To prepare gfp-bcl-X L-contained recombinant adenovirus(rAd-gfp-bcl-X L).METHODS: Bcl-X L gene was amplified from pEGFP-C 3-bcl-X L, subcloned into shuttle plasmid and formed transfer plasmid of pAdTrack-CMV-bcl-X L. Then pAdTrack-CMV-bcl-X L was linealinzed with PmeI and co-transformed into BJ5183 bacteria with adenovirus genomic plasmid of pAdEasy-1. The identified recombinant adenovirus plasmid was digested with PacI and transfected into 293 cells to package recombinant adenovirus particles. The target gene was detected by PCR.RESULTS: There were about 35% positive recombinant bacterial clones after the co-transformation of pAdTrack-CMV-bcl-X L and pAdEasy-1 into BJ5183. Recombinant adenovirus particle were produced and further amplified after the transfection of pAdEasy-1-gfp-bcl-X L into 293 cells. PCR test indicated that the recombinant Ad contained bcl-X L gene. The titer of the purified rAd-gfp-bcl-X L was 6 5?10 12 PFU/L. CONCLUSIONS: The homologous recombination in bacteria is a convenient and high efficient method to prepare rAd-gfp-bcl-X L. This affords a good gene transfer vector for the gene therapy in human's diseases.

The expression of NF-?Bp65 and bcl-x_ L gene in pancreatic carcinoma and their significance / 中国现代普通外科进展

Objective:To study the expression of NF-?Bp65 and bcl-x L gene in pancreatic carcinoma and their clinical significance.Methods:The expressions of NF-?Bp65 and bcl-x L protein in 52 pancreatic carcinoma were studied by immunohistochemistry,and their relationship to clinical pathology were analyzed.Results:The positive rates of NF-?Bp65 protein and bcl-x L protein in pancreatic carcinoma were 63.5% and 73.1% respectively,and which were higher than those of normal pancreatic tissues.The expression of NF-?Bp65 was related to tumor diameter,TNM stage and lymph node metastasis,and had no relation with the extent of differentiation and type of pathology.There was positive correlation between NF-? Bp65 and bcl-x L expression.Conclusion:NF-? Bp65 may be an important oncoprotein,and may have positive regulation on bcl-x L,then promote tumorgenesis.

Relationship between the Expression of Apoptosis-Related Proteins and Chemosensitivity in Gastric Cancer Cell Lines

BACKGROUND: There has been a growing realization that a variety of anticancer drugs can induce apoptotic cell death. In the present study, an attempt was made to investigate the responsiveness of gastric cancer cells to various anticancer drugs and to identify which apoptosis-related proteins could be correlated to chemosensitivity. METHODS: Nine human Korean gastric cancer cell lines (SNU-1, -5, -16, -484, -601, -620, -638, -668, and -719) were analyzed. The cytotoxicity of each cell line to camptothecin, cisplatin, mitomycin C, vincristine, 5-FU, epirubicin, and doxorubicin was determined by using a MTT (dimethylthiazole- diphenyltetrazolium-bromide) assay. Apoptosis-related proteins (p53, p21, Bcl-2, Bcl-x, and Bax) were detected using a Western blot assay. RESULTS: Of the nine gastric cancer cell lines, SNU-1 was resistant while SNU-5 was sensitive to anticancer drugs. Mutated p53 was detected in all the cell lines. The highest expression of Bcl-2 was observed in SNU-1 while less or no expression of Bcl-2 was observed in SNU-5, -484, and -601. Bcl-xL was less expressed in SNU-5 than in the other cell lines. CONCLUSIONS: Chemosensitivity in gastric cancer cell lines was correlated mainly with the level of Bcl-2 and partly with that of Bcl-xL. There was no correlation between the chemosensitivity and other apoptosis-related proteins, such as p21, p53, Bax, and Bcl-xS in the studied gastric cancer cell lines.