Positive Expression Of Bcl-2 Research Articles

Promethazine inhibits neuronal apoptosis via PI3K/Akt signaling pathway in rats with cerebral infarction.

To study the effect of promethazine on neuronal apoptosis in rats with cerebral infarction (CI) through the phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/Akt) signaling pathway. A total of 36 Sprague-Dawley rats were randomly divided into the sham group (n=12), model group (n=12), and promethazine group (n=12). The external carotid artery was only exposed in the model group, and the ischemia-reperfusion model after CI was established using the suture method in the other two groups. After modeling, the normal saline was intraperitoneally injected in the sham group and model group, while promethazine was intraperitoneally injected in the promethazine group. The rats were sampled after 1 week of intervention. The neurological deficits of rats were evaluated using the Zea-Longa score, and the cognitive function, the spatial learning, and memory of rats were detected via the water maze test. Moreover, the expressions of B-cell lymphoma-2 (Bcl-2) and Bcl-2 associated X protein (Bax) in brain tissues were detected via immunohistochemistry, and the relative protein expressions of PI3K p85, PI3K p110, and p-Akt were detected via Western blotting. The mRNA expressions of Bax and Bcl-2 were detected via quantitative Polymerase Chain Reaction (qPCR), and the apoptosis was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay. The Zea-Longa score was significantly increased in the model group and promethazine group compared with that in the sham group (p<0.05), while it significantly declined in the promethazine group compared with that in the model group (p<0.05). The escape latency was significantly prolonged and the times of crossing platform were significantly reduced in the model group and promethazine group compared with those in the sham group (p<0.05), while the escape latency was significantly shortened and the times of crossing platform were significantly increased in the promethazine group compared with those in the model group (p<0.05). Compared with those in the sham group, the positive expression of Bax was significantly increased, while the positive expression of Bcl-2 was remarkably decreased in the model group and promethazine group (p<0.05). Compared with those in the model group, the positive expression of Bax was significantly decreased, while the positive expression of Bcl-2 was remarkably increased in the promethazine group (p<0.05). Besides, the model group and promethazine group had evidently higher relative protein expressions of PI3K p85, PI3K p110, and p-Akt than the sham group (p<0.05), while the promethazine group also had evidently higher relative protein expressions of PI3K p85, PI3K p110, and p-Akt than the model group (p<0.05). Compared with the sham group, model group, and promethazine group had remarkably increased relative mRNA expression of Bax, and remarkably decreased relative mRNA expression of Bcl-2 (p<0.05). Compared with those in the model group, the relative mRNA expression of Bax was remarkably decreased, while the relative mRNA expression of Bcl-2 was remarkably increased in the promethazine group (p<0.05). Finally, the apoptosis rate was significantly higher in the model group and promethazine group than that in the sham group (p<0.05), while it was significantly lower in the promethazine group than that in the model group (p<0.05). Promethazine inhibits neuronal apoptosis in CI rats by upregulating the PI3K/Akt signaling pathway, thereby exerting a protective effect.

Melatonin inhibits the inflammation and apoptosis in rats with diabetic retinopathy via MAPK pathway.

To investigate the effect of melatonin on diabetic retinopathy rats through the mitogen-activated protein kinase (MAPK) pathway. A total of 48 Sprague Dawley (SD) rats were randomly divided into normal group (n=12), model group (n=12), melatonin group (n=12), and inhibitor group (n=12). The rats in normal group received no treatment. Those in model group, melatonin group, and inhibitor group were prepared into models of diabetic retinopathy and intraperitoneally injected with normal saline, melatonin, and SB 203580, respectively. After 7 days of intervention, the materials were taken. The expressions of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) were detected through immunohistochemistry. Western blotting was employed to determine the protein expression levels of p38 MAPK, phosphorylated (p)-p38 MAPK, and cysteinyl aspartate specific proteinase-3 (Caspase-3). The messenger ribonucleic acid (mRNA) expression levels of Bax and Bcl-2 were measured via quantitative Polymerase Chain Reaction (qPCR). Enzyme-linked immunosorbent assay (ELISA) was performed to detect the levels of serum interleukin-1β (IL-1β) and IL-18. The apoptosis was determined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling (TUNEL). Based on immunohistochemistry, model group, melatonin group, and inhibitor group exhibited significantly increased positive expression of Bax but notably decreased positive expression of Bcl-2 in comparison with normal group (p<0.05). Compared with those in model group, the positive expression of Bax was clearly reduced, while the positive expression of Bcl-2 was overtly raised in melatonin group and inhibitor group (p<0.05). The results of Western blotting showed that there was no difference in the protein expression of p38 MAPK among all groups (p>0.05). Compared with normal group, the other three groups had remarkably elevated protein expressions of p-p38 MAPK and Caspase-3 (p<0.05). The protein expressions of p-p38 MAPK and Caspase-3 in melatonin group and inhibitor group were significantly lower than those in model group decreased (p<0.05). QPCR assay revealed that the mRNA expression of Bax was markedly lower in normal group than that in the other three groups, while the mRNA expression of Bcl-2 was significantly higher in normal group than that in the other three groups (p<0.05). Compared with model group, melatonin group, and inhibitory group showed clearly declined mRNA expression level of Bax and notably increased mRNA expression level of Bcl-2 (p<0.05). TUNEL results revealed that the apoptosis rate was remarkably elevated in the other three groups compared with that in normal group (p<0.05). In comparison with model group, melatonin group and inhibitor group exhibited significantly reduced apoptosis rate (p<0.05). Melatonin inhibits the inflammation and apoptosis in rats with diabetic retinopathy by repressing the MAPK pathway.

Effect of propofol on myocardial ischemia/reperfusion injury in rats through JAK/STAT signaling pathway.

The aim of this study was to investigate the influences of propofol on myocardial ischemia/reperfusion injury in rats through the Janus kinase/signal transducers and the activators of transcription (JAK/STAT) signaling pathway. A total of 48 Sprague-Dawley (SD) rats were randomly divided into four groups, including: the sham-operation group (n=12), the model group (n=12), the propofol group (n=12) and the inhibitor group (n=12). The rats in the sham-operation group only received thoracotomy, without the modeling of the ischemia/reperfusion injury. The model of myocardial ischemia/reperfusion injury was established in the rats of the model group, and the rats were given normal saline for intervention. The rats in the propofol group were utilized to prepare the model of myocardial ischemia/reperfusion injury and were intervened with propofol. Meanwhile, the rats in the inhibitor group received intervention with AG490 after the establishment of myocardial ischemia/reperfusion injury model. Immunohistochemistry was applied to detect the expressions of B-cell lymphoma-2 (Bcl-2) and Bcl-2-associated X protein (Bax). Western blotting was utilized to measure the relative protein expressions of phosphorylated JAK2 (p-JAK2) and p-STAT3. The messenger ribonucleic acid (mRNA) expressions of Bax and Bcl-2 were determined via quantitative Polymerase Chain Reaction (qPCR). Furthermore, cell apoptosis was examined using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay. Immunohistochemistry results showed that compared with the sham-operation group, the positive expression of Bax remarkably increased (p<0.05), while Bcl-2 notably decreased (p<0.05) in the model group, propofol group, and inhibitor group. The propofol group and inhibitor group showed a significant lower positive expression of Bax (p<0.05) and evident higher positive expression of Bcl-2 (p<0.05) when compared with the model group. However, there were no significant differences in the positive expressions of Bax and Bcl-2 between the propofol group and inhibitor group (p>0.05). According to the results of Western blotting, the relative protein expression levels of p-JAK2 and p-STAT3 proteins were remarkably elevated in the model group, propofol group and inhibitor group in comparison with those in the sham-operation group (p<0.05). Propofol group and inhibitor group exhibited remarkably lower protein expression levels of p-JAK2 and p-STAT3 compared with the model group (p<0.05). However, no significant differences were observed in the protein expressions of p-JAK2 and p-STAT3 between propofol group and inhibitor group (p>0.05). The results of qPCR manifested that the mRNA expression of Bax was notably higher (p<0.05), whereas Bcl-2 was significantly lower (p<0.05) in the model group, propofol group and inhibitor group than those of the sham-operation group. Compared with the model group, the mRNA expression of Bax was evidently declined (p<0.05), while Bcl-2 was significantly elevated (p<0.05) in the propofol group and inhibitor group. Meanwhile, there were no evident differences in the mRNA expressions of Bax and Bcl-2 between the propofol group and inhibitor group (p>0.05). Subsequent TUNEL assay indicated that the model group, propofol group, and inhibitor group showed remarkably higher apoptosis rate than the sham-operation group (p<0.05). Moreover, the apoptosis rate was remarkably reduced in the propofol group and inhibitor group in comparison with the model group (p<0.05). However, no significant difference was observed in the apoptosis rate between propofol group and inhibitor group (p>0.05). Propofol inhibits myocardial cell apoptosis after myocardial ischemia/reperfusion injury by repressing the JAK/STAT signaling pathway.

LncRNA LINC00668 promotes the progression of breast cancer by inhibiting apoptosis and accelerating cell cycle.

Objective: To elucidate how lncRNA 00668 (LINC00668) influences the development of breast cancer (BC).Materials and methods: Genome-wide expression profile of BC and paracancerous tissues were downloaded from The Cancer Genome Atlas (TCGA) and BC tissues and paracancerous tissues enrolled from our hospital for analyzing the expression level of LINC00668 and its correlation with prognosis. GSEA was conducted to analyze the potential functions of LINC00668. By transfection of sh-LINC00668 in BC cells, proliferation, apoptosis, cell cycle and colony formation of BC cells were accessed. Western blot was conducted to detect protein expressions of Ki-67, CDK4, Bcl-2, p21 and genes in AKT/mTOR pathways after LINC00668 knockdown in BC cells. Finally, tumor-bearing nude mice were administrated with BC cells. We compared the proliferative rate in mice with different administrations. Immunohistochemistry was carried out to access expression levels of Ki-67, CDK4, Bcl-2 and P21 in mice.Results: Both TCGA data and BC tissues harvested from our hospital indicated the higher expression of LINC00668 in BC tissues. LINC00668 expression was negatively correlated to prognosis of BC patients. GSEA pointed out that LINC00668 is enriched in regulations of cell cycle and apoptosis. By transfection of sh-LINC00668 in MDA-MB-231 and MDA-MB-436 cells, the proliferative and colony formation abilities of BC cells decreased. Besides, LINC00668 knockdown in BC cells induced apoptosis and arrested cell cycle. LINC00668 knockdown downregulated Ki-67, CDK4 and Bcl-2, but upregulated p21. The AKT/mTOR pathway was inhibited after LINC00668 silenced. In vivo experiments demonstrated the decreased proliferative rate in tumor-bearing mice administrated with sh-LINC00668 transfected BC cells. Consistently, immunohistochemical results showed lower positive expressions of Ki-67, CDK4 and Bcl-2, but higher positive expression of p21 in sh-LINC00668 group.Conclusion: LINC00668 is highly expressed in BC tissues and can promote the progression of BC by inhibiting apoptosis and accelerating cell cycle progression.

MiR-30a-5p modulates traits of cutaneous squamous cell carcinoma (cSCC) via forkhead box protein G1 (FOXG1).

miRNA has shown its potential in the regulation of squamous cell carcinoma (SCC). However, the mechanism of such an effect was not quite clear. Therefore, we aimed to investigate whether miR-30a-5p participated in the regulation of cutaneous SCC (cSCC) and the possible mechanism involved. 5-Ethynyl-2'-deoxyuridine (EdU) and cell cycle were measured using flow cytometry. The formation of cell colony was tested by colony formation assay. The capacities of migration and invasion were tested by wound healing assay and Transwell invasion assay, respectively. The target of miR-30a-5p was predicted by bioinformatics and identified by luciferase assay. Western blot was used for the determination of proteins and qPCR was for mRNA levels. miR-30a-5p expression was lowered in SCL-1 and A431 cells, and its upregulation suppressed EdU positive cells, colony numbers, migration, invasion and Bcl-2 expression, and elevated Bcl-2-associated X protein (Bax) and cleaved Caspase-3 expressions, arresting cell cycle in G1 phase. Moreover, forkhead box protein G1 (FOXG1) was proved to be the target of miR-30a-5p, and FOXG1 overexpression partially offsets the decreased colony numbers, migration and invasion rates due to miR-30a-5p overexpression in SCL-1 and A431 cells. miR-30a-5p showed a regulatory role on the expression of FOXG1 and further modulated the progressing of cSCC cells, which could be a novel pathway intervening the development of cSCC.

Comparison of BCL-2 Ecpression in Periocular Basal Cell Carcinoma Aggressive and Non-Aggressive Types in Mohammad Hoesin Hospital Palembang

Abstract&#x0D; Background: Basal cell carcinoma (BCC) is a malignant tumor originating from non- keratinizing cells originating from the basal layer of the epidermis. Basal cell carcinoma is the most common malignant tumor, nearly 90% of all non-melanoma skin cancers. Clinically, BCC is difficult to distinguish, final diagnosis determined based on histopathological examination of complete tumor tissue. Based on histological growth patterns, BCC is divided into two, non- aggressive type (nodular and superficial) and aggressive type (infiltration, micronodular, fibroepithelial, basoskuamosa, keratotic, adnexal differentiation). The examination technique by analyzing the proteins or genes involved is related to the aggressiveness of basal cell carcinoma.&#x0D; Objective: This study was conducted to determine the comparison of expression of Bcl-2 in periocular basal type aggressive and non-aggressive carcinomas in dr. Mohammad Hoesin Palembang.&#x0D; Methodology: A cross sectional analytic study from 26 periocular basal cell carcinoma patients in RSUP dr. Mohammad Hoesin Palembang, January 2014- June 2018. Histopathologic features were examined with hematoxylin-eosine staining and imunohistochemical were examined with Bcl-2 monoclonal antibody. Data were analyzed by bivariate analysis, using chi-square test and fisher exact test.&#x0D; Results: Positive Bcl-2 expression was more common in periocular basal cell carcinoma (93.8%) than aggressive (50%), and nodular type (93.8%) compared to other types of histological differentiation. Positive Bcl-2 expression was also higher in female patients (90.9%) than males (66.7%). There is a significant relationship between expression of Bcl-2 and growth patterns, differentiation of cell histology and also sex of patients with basal cell carcinoma.&#x0D; Conclusion: In this study, there was a significant correlation between expression of Bcl-2 and aggressive and non-aggressive periocular basal cell carcinoma in RSUP dr. Mohammad Hoesin Palembang.

Expresión de p53 y Bcl-2 en carcinoma ductal infiltrante de mama localmente avanzado. Su relación con el subtipo intrínseco molecular como factor pronóstico

In breast cancer there are prognostic and predictive factors that can affect the evolution of women with breast cancer. For this reason, the present study aims to determine the immunohistochemical expression (IHC) of p53 and Bcl-2 proteins in the primary tumor of 285 women with a diagnosis of infiltrating ductal carcinoma of the breast under follow-up at the Dr. Miguel Oncology Institute. Perez Carreno from Valencia, Venezuela. The immunoexpres-sion of both proteins was related to the molecular intrinsic subtypes of breast cancer defined by IHC: Luminal A, Luminal B, HER2 and Triple Negative (TN), disease free survival (SLE) and overall survival (OS). The tumors p53 negative and Bcl-2 positive showed a higher SLE and SG. In the p53 negative tumors of the Luminal A subtype there was a higher SLE (p = 0.005), although in the Luminal tumors (A and B) the OS was higher with p <0.05. In addition, greater SLE was presented in Luminal and TN cases with positive Bcl-2 expression, but without statistical significance. On the contrary, there was a significant prognostic effect of the expression of Bcl-2 in the TN subtype, with a lower mean SG compared to the SG obtained in tumors with positive Bcl-2. Finally, based on the findings found, the absence of p53 expression and the positive expression of Bcl-2 are good prognostic factors, with statistical significance in Luminal and TN tumors, respectively.

Suppressed microRNA-96 inhibits iNOS expression and dopaminergic neuron apoptosis through inactivating the MAPK signaling pathway by targeting CACNG5 in mice with Parkinson\u2019s disease

BackgroundThere have been a number of reports implicating the association of microRNAs (miRs) and the MAPK signaling pathway with the dopaminergic neuron, which is involved in the development of Parkinson’s disease (PD). The present study was conducted with aims of exploring the role of miR-96 in the activation of iNOS and apoptosis of dopaminergic neuron through the MAPK signaling pathway in mice with PD.MethodsThe miR and the differentially expressed gene in PD were screened out and the relationship between them was verified. A mouse model of PD induced by MPTP and was then constructed and treated with miR-96 mimic/inhibitor and CACNG5 overexpression plasmid to extract nigral dopaminergic neuron for the purpose of detecting the effect of miR-96 on PD. The TH and iNOS positive neuronal cells, the apoptotic neuronal cells by TUNEL staining, and expression of miR-96, CACNG5, iNOS, p38MAPK, p-p38MAPK, c-Fos, Bax, and Bcl-2 in substantia nigra dopaminergic neuronal tissues were evaluated.ResultsThe results obtained from the aforementioned procedure were then verified by cell culture of the SH-SY5Y cells, followed by treatment with miR-96 mimic/inhibitor, CACNG5 overexpression plasmid and the inhibitor of the MAPK signaling pathway. CACNG5 was confirmed as a target gene of miR-96. The inhibition of miR-96 resulted in a substantial increase in nigral cells, TH positive cells and expression of CACNG5 and Bcl-2 in nigral dopaminergic neuronal tissues, and a decrease in iNOS positive cells, apoptotic neuronal cells, and expression of iNOS, p38MAPK, p-p38MAPK, c-Fos, and Bax.ConclusionThe above results implicated that the downregulation of miR-96 inhibits the activation of iNOS and apoptosis of dopaminergic neuron through the blockade of the MAPK signaling pathway by promoting CACNG5 in mice with PD.

Protein expressions of AIB1, p53 and Bcl-2 in epithelial ovarian cancer and their correlations with the clinical pathological features and prognosis.

To investigate the protein expressions of steroid receptor coactivator amplified in breast cancer 1 (AIB1), apoptosis-related protein p53 and B-cell lymphoma-2 (Bcl-2) in epithelial ovarian cancer; to analyze the correlations among the expressions of these three proteins; to explore their correlations with the clinical pathological features and prognosis. Immunohistochemistry streptavidin-peroxidase (IHC-SP) method was performed to detect the positive protein expressions of AIB1, Bcl-2, and p53 in the pathological sections of normal ovarian tissues, benign ovarian epithelial tumor, and epithelial ovarian cancer, thereby analyzing the protein expression rates of these genes in different pathological stages, lymphatic metastasis and postoperative recurrent ovarian cancer, and carrying out the correlation analysis of these three proteins. Positive protein expressions of AIB1, Bcl-2, and p53 were identified in the epithelial ovarian cancer tissues, and with an increase in the tumor staging of ovarian cancer, we found that the positive protein expression rate was gradually augmented. Particularly in ovarian cancer with lymphatic metastasis and postoperative recurrence, the positive expression rate was almost 100%. The protein expression of AIB1 was positively correlated with those of the p53 and Bcl-2. We showed that positive expressions of AIB1, Bcl-2, and p53 in ovarian cancer are closely correlated with the pathological staging, metastasis, and recurrence, and the positive protein expression suggests a poor prognosis.

Experimental study of urinary center change in pons after conus medullaris injury in rats

To observe the structural changes of urinary center and the expression of Bcl-2 after conus medullaris injury in rats brain so as to explore the possible influence factors of degeneration in brain. Thirty-six adult Sprague-Dawley rats were randomly divided into experimental group ( n=30) and control group ( n=6). In the experimental group, the conus medullaris injury model was established by cutting off the spinal nerve below L 4, and no treatment was done in the control group. The modeling operations in the experimental group were successful, and 2 rats died at 3 months and 5 months after modeling operation respectively, which may be caused by renal failure or urinary tract infection. In the experimental group, 6, 6, 6, 5, and 5 rats were killed at 1 day, 1 week, and 1, 3, 6 months after operation respectively, and 1 rat was killed at each time point in the control group. The dorsolateral tissue of the pontine tegmentum was harvested to perform HE staining and Bcl-2 immunohistochemical SP staining. HE staining showed that there was no obvious difference between the experimental group and the control group at 1 day after operation, the neurons were densely packed, arranged neatly, and the nucleoli were clear; at 1 week, the space between the neurons in the experimental group were slightly widened; at 1 month, nucleus retraction in some neurons happened in the experimental group; at 3 and 6 months, the nuclei in the experimental group were more and more condensed, and even some cells disappeared. Bcl-2 immunohistochemical SP staining showed that the expression of Bcl-2 in the control group was weakly positive. The positive expression of Bcl-2 was found at 1 day after operation in the experimental group; the positive expression of Bcl-2 at 7 days after operation was significantly higher than that in the control group, and reached the peak; the positive expression of Bcl-2 decreased gradually at 1, 3, and 6 months after modeling operation, but it was still higher than that of the control group. The urinary center appears structure degeneration and necrocytosis after conus medullaris injury in rats brain. The elevated expression of Bcl-2 may be associated with brain tissue repair and function remodeling.

Distinct immunophenotypes and prognostic factors in renal cell carcinoma with sarcomatoid differentiation: a systematic study of 19 immunohistochemical markers in 42 cases

BackgroundRenal cell carcinoma (RCC) with sarcomatoid differentiation is a relatively rare tumor containing both carcinoma and sarcomatoid components. However, there has not been a systemic study on immunophenotypes of renal cell carcinoma with sarcomatoid differentiation, especially using some renal specific immunohistochemical markers. In this study, we aimed to comprehensively investigate the distinct immunophenotypes of RCC with sarcomatoid differentiation to analyze the pathogenesis of sarcomatoid differentiation and identify new prognostic factors in RCC with sarcomatoid differentiation.MethodsA total of 42 cases of RCCs with sarcomatoid differentiation were enrolled into the study. Immunohistochemistry study was performed on tissue microarrays to evaluate the expressions of 19 immunohistochemical markers including a series of epithelial, mesenchymal markers and RCC specific markers. Kaplan-Meier method was applied to assess the prognostic values of CD10, CAIX, p53 and Bcl-2.ResultsHistologically, 42 cases of RCCs with sarcomatoid differentiation presented with different proportions of carcinoma and sarcomatoid components. The cohort contained 35 cases of clear cell renal cell carcinoma (CCRCC) and 7 cases of chromophobe renal cell carcinoma (ChRCC) based on the carcinoma components. Immunohistochemically, all cases were positive for vimentin, and 80% of cases showed immunostaining for at least one epithelial marker, such as CK, EMA, CK7 and CK18. Notably, the expression rates of CAIX, CD10 and PAX8 in sarcomatoid cells were 76%, 76% and 64%, respectively. The carcinoma component of the tumors showed differentient labeling for CAIX, CD10, vimentin, CK7 and CD117 in CCRCC vs ChRCC, but the sarcomatoid component lost the specificity for these markers (p < 0.05). Patients with positive expressions of CAIX, p53 and Bcl-2 had a poor prognosis.ConclusionsThe sarcomatoid cells in RCC with sarcomatoid differentiation express both epithelial and mesenchymal markers, supporting their epithelial origin. PAX8, CAIX and CD10 could be used as the reliable and useful markers to determine the renal origin of sarcomatoid cells such as in fine needle aspiration cases and metastatic RCC with sarcomatoid differentiation. CAIX, p53 and Bcl-2 might play important roles in the transformation from renal cell carcinoma to high malignant sarcomatoid differentiation, and these three immunohistochemical markers are adverse prognostic factors for the survival of patients with RCC with sarcomatoid differentiation.

Effects of ropivacaine-induced convulsion on cell apoptosis of hippocampus and the ability of learning and memory in adult rats

Objective To investigate the influence of ropivacaine-induced convulsion on cell apoptosis of hippocampus and the ability of learning and memory in adult rats. Methods One hundred and ten SD adult rats were randomly divided into two groups: group NS (n=35) and group R (n=75) . Group R were i.p injected of convulsion ED50 of ropivacaine and rats undergoing convulsion were selected for experiment. Group NS were i.p. injected of NS. At 2 h, 6 h, 24 h, 3 d, 7 d after convulsion, 5 rats in each group were chosen, then their hippocampus tissues were embedded with paraffin and siced up for immunohistochemistry experiments. The positive expressions of Bax and Bcl-2 in hippocampus were calculated with microscope. Ten rats in each group were chosen and the effects of ropivacaine-induced convulsion on spatial memory and learning ability were observed in 8 day and 45 day after convulsion. Results Convulsion latency and persistence time of group R were (9.3±3.7) s and (7.2±4.6) s. The expression of Bax of group R increased at 2 h after convulsion, reached the top at 24 h and then declined, which was still higher than group NS in 7 d. The expression of Bcl-2 of group R increased at 6 h (P 0.05) . Conclusions Convulsion induced by i.p. convulsion ED50 of ropivacaine increased cells apoptosis of hippocampus and reduced the ability of learning and memory in earlier period after convulsion, but it had little impact in longer period after convulsion. Key words: Ropivacaine; Convulsion; Apoptosis; Learning; Memory; Rat

The complex morphologic diagnostic of marginal zone lymphoma.

The purpose of study: To determine possibilities of complex morphological diagnostic (routine cytology and histology, immunocytochemistry and immunohistochemistry, molecular genetics) of lymphoma of marginal zone. The study included 10 patients with diagnosis of lymphoma of marginal zone cells, established on the basis of application of morphological (cytological and histological), immunomorphologic (immunocytochemical and immunohistochemical) and molecular genetic techniques. The immunofenotyping was implemented using immunocytochemical technique (EnVision FLEX) applying monoclonal antibodies by DAKO manufacturer. The immunofenotyping was implemented by flow cytofluorometry technique (flow cytofluorometer FACS Calibur by Becton Dikinson, USA) using antibodies by DACO manufacturer labeled by fluorescent marks (FITC or RPE). The antibody panel included: common leukocytic antigen, common cytokeratins CD19, CD20, CD79a,CD10, Bcl2, Bcl6, CD23, CD34, TdT, CD3, CD4, CD5, CD8, CyclinD1, Ki67, κ, λ. The FISH technique was applied using probes Bcl2 FISH DNA Probe, Split Signal и MALT1 FISH DNA Probe Split Signal by DAKO manufacturer. In one patient a gene Bcl2 change was detected with purpose of differentiating diagnostic with follicular lymphoma. In three patients, diagnosing lymphoma of marginal zone was implemented by FISH technique using detection of translocation (11;18)(q21;q21), involving gene of inhibitor of apoptosis API2 and gene MALT1. The routine cytological analysis of ten cases permitted to establish an exact diagnosis of lymphoma only in five cases and with no indication that it is lymphoma of marginal zone. In two cases under routine cytological analysis only a suspicion about lymphoma was suggested. In all ten patients a positive expression of pan-B-cellular markers CD19, CD20, CD79a was observed both under immunocytochemistry and immunohistochemistry and flow cytofluorometry. In 6 patients (60%) a positive expression of gene Bcl2 was observed both under immunocytochemistry and immunohistochemistry and flow cytofluorometry. The comparison of immunocytochemistry and immunohistochemistry and flow cytofluorometry established absence of expression of CD5, CD3, CD10, CD34, CD23, Bcl6, TdT, cyclin D1. Under immunocytochemistry and immunohistochemistry in ten cases proliferative activity protein Ki-67made up no more than 30%. Under flow cytofluorometry clonality on light chains of immunoglobulins κ or λ (Igλ/Igκ) were established. Overall, correlation coefficient (r, p < 0,05) between immunocytochemistry and immunohistochemistry, flow cytofluorometry made up to 1. The four patients were applied FISH-reaction for adjustment of diagnosis. In patient with nodal lymphoma of marginal zone gene Bcl2 change was absent. In three patients with MALT-lymphoma a gene MALT1 change was established. Thereby, accuracy of routine cytological analysis at diagnosing lymphoma without indication of its type in case of lymphoma of marginal zone made up to 50%, sensitivity - 50%, specificity - 100%. The accuracy of immunofenotyping permitting diagnosing lymphoma of marginal zone made up to 100%, sensitivity - 100%, specificity - 100%. The correlation coefficient (r, p < 0,05) between data of immunocytochemistry and immunohistochemistry, flow cytofluorometry made up to 1. The accuracy, sensitivity and specificity of complex analysis (cytology, immunocytochemistry and FISH-technique) in diagnostic of lymphoma of marginal zone made up to 100%.

Identification of prognostic factors in patients with diffuse large B-cell lymphoma.

The correlation between various clinicopathological factors and 5-year survival rate, the correlation between those factors with the International Prognostic Index, the concordance between Hans' and Choi's approach was evaluated. The survival in different subtypes as classified by Hans' or Choi's approach was mapped. The Eastern Cooperative Oncology Group (ECOG) performance score 2-5, positive Bcl-2 expression, negative CD10 expression or negative Bcl-6 expression significantly correlated with worse prognosis. The two algorithms showed good consistency (83% concordance, Kappa = 0.660, P < 0.001). By both classifications, the 5-year overall survival rate in germinal center B-cell-like subtype (GCB) lymphoma is significantly higher than that in the non-GCB subtype. There were 25 cases assigned to conflicting subtypes by the two approaches. Among these 25 cases, ECOG 2-5, positive Bcl-2 expression, negative CD10 expression, or negative Bcl-6 expression significantly correlated with worse prognosis. ECOG 2-5, positive Bcl-2 expression, negative CD10 expression, or negative Bcl-6 expression are independent markers for poor prognosis of DLBCL patients. There were 15% cases assigned to conflicting subgroups based on the two algorithms. For these cases, ECOG 2-5, positive Bcl-2 expression, negative CD10 expression, or negative Bcl-6 expression still significantly correlate with poor prognosis.

Folic acid ameliorates L-thyroxin induced hepatotoxicity and oxidative stress in albino rats.

Thyroid hormones have been known to regulate the energy metabolism of most tissues including liver. Alterations in their normal levels cause some biochemical and clinical abnormalities such as hypothyroidism and hyperthyroidism. The present study evaluated the effect of thyroid hormone, L-thyroxin on liver of albino rats. Additionally the ameliorating role of folic acid supplementation was investigated. Fifty male albino rats were randomly divided into five groups (group I, control; group II, folic acid; group III, L-thyroxin sodium administration (100 μg/kg / body weight); group IV, L-thyroxin and folic acid group and V, recovery group). The results showed that there were a significant increase in ALT, AST, MDA and nitric oxide in L-thyroxin treated rats as compared to control group. On the other hand, a significant decrease in glutathione (GSH) in L-thyroxin treated rats as compared to control group. Histological results showed that liver sections of L-thyroxin group showed histopathological lesions such as leucocytic infiltrations, congestion of central and portal veins and cytoplasmic vacuolation of hepatocytes with the presence of pyknotic nuclei, in addition to fatty infiltration. Immunohistochemical results revealed that strong positive expression of PCNA, P53, and Bcl-2 were detected in the liver section in L-thyroxin treated rats and recovered rats as compared to control and folic acid groups. However; mild to moderate positive expressions of PCNA, P53, and Bcl-2 were observed in rats treated with L-thyroxin and folic acid in liver section. This reflects oxidative stress associated with hyperthyroid state.

The study of traditional Chinese medical elongated-needle therapy promoting neurological recovery mechanism after spinal cord injury in rats

Ethnopharmacological relevanceAcupuncture is a key part of traditional Chinese medicine, shown to induce favorable neuroplasticity for injuries in the central and peripheral nervous systems. Recent studies report elongated needle therapy (ENT) with BL54 and ST28 may restore acute spinal cord injury (ASCI). However, the precise mechanism for this has not been elucidated. Aim of the studyIn our current study, we investigated the effects of ENT on inflammation and neuronal apoptosis induced by ASCI, and whether PI3K/Akt and MAPK/ERK signaling pathways are involved in the ENT restoration effect. Materials and methodsRat models of moderate SCI were established in accordance with the modified Allen's method and were treated with ENT continuously for 7 days. Spontaneous activities were evaluated by the Basso Beattie and Bresnahan locomotor scale. Levels of inflammatory cytokines, such as tumor necrosis factor alpha, interleukin-6, IL-1β, and nuclear factor kappa-β, were determined by enzyme-linked immunosorbent assay. Cell apoptosis was examined by the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. The proportions of cells with positive Bcl-2 and Bax expression were determined by immunohistochemical assays, whilst the expression profiles of p-AKT and p-ERK in spinal cord tissues were evaluated by western blotting. Furthermore, the expression profiles of Cytochrome-C (Cyt-C) and caspase-3 in vivo were analyzed by reverse transcription polymerase chain reaction. The potential inhibitory effects downstream of the Akt and ERK signaling pathways were examined by administration of specific inhibitors LY294002 and PD98059 in vivo. ResultsAs indicated by this study, inflammation as well as PI3K/Akt- and MAPK/ERK signaling pathway-mediated neuronal apoptosis were involved in the course of SCI in rats. The neuro-protective effect of ENT was associated with reduced Bax protein-positive neurons and increased Bcl-2 protein-positive neurons. ENT enhanced recovery of rat activities. Activation of p-Akt and p-ERK in the PI3K/Akt and MAPK/ERK signaling pathways, inhibited expression of the critical component Cyt-C. Cyt-C is required for the mitochondrial apoptosis pathway and cascade of caspase-3, which is involved in activation of neuronal apoptosis through down-regulation of Bax protein and up-regulation of Bcl-2, as determined by TUNEL. The administration of PI3K/Akt and MAPK/ERK signaling pathway specific inhibitors, LY294002 and PD98059, suppressed expression of both p-Akt and p-ERK. ConclusionENT with BL54 and ST28 points can promote the recovery of ASCI. And the neuro-protective effect of ENT during the restoration of SCI may be associated with the suppression of both inflammation and activation of PI3K/Akt and MAPK/ERK signaling pathways, resulting from down-regulation of Bax protein, up-regulation of Bcl-2, and inhibition of the mitochondrial apoptosis pathway.

BCL2 as a Subtype-Specific Prognostic Marker for Breast Cancer.

PurposeB-cell lymphoma 2 (BCL2) is an antiapoptosis protein and an important clinical breast cancer prognostic marker. As the role of BCL2 is dependent on the estrogen receptor (ER) status, this effect might differ according to molecular subtypes. The aim of this study was to evaluate the relationship between the prognostic outcomes and BCL2 expression among the molecular subtypes.MethodsWe retrieved the data of 1,356 patients who were newly diagnosed with malignant breast cancer between November 2006 and November 2011. Immunohistochemistry was used to measure ER, progesterone receptor, human epidermal growth factor receptor 2 (HER2), Ki-67, and BCL2 expression. We classified breast cancer into five molecular subtypes based on the 13th St. Gallen International Expert Consensus, including luminal A, luminal B (HER2-negative), luminal B (HER2-positive), HER2-overexpression, and triple-negative subtypes. We analyzed the clinicopathological features and assessed the correlation between BCL2 expression and clinical outcomes, such as relapse-free survival (RFS) and disease-specific survival (DSS) according to the five molecular subtypes.ResultsA total of 605 cases of breast cancer (53.8%) showed BCL2 expression. BCL2-positive expression was associated with young age (<50 years, p=0.036), lower histological grade (p<0.001), low Ki-67 level (<14%, p<0.001), hormone receptor positivity (p<0.001), HER2 negativity (p<0.001), luminal breast cancer (p<0.001), and low recurrence rate (p=0.016). BCL2-positive expression was also associated with favorable 5-year RFS (p=0.008, 91.4%) and DSS (p=0.036, 95.6%) in all the patients. BCL2-positive expression in luminal A breast cancer resulted in significantly favorable 5-year RFS and DSS (p=0.023 and p=0.041, respectively). However, BCL2 expression was not associated with the prognosis in the other subtypes.ConclusionThe prognostic role of BCL2 expression in breast cancer is subtype-specific. BCL2 expression differs according to the molecular subtype and is a good prognostic marker for only luminal A breast cancer.

Clinicopathological correlation of Bcl-2 oncoprotein expression in oral precancer and cancer

Oral squamous cell carcinoma is the most common malignant tumor of the oral cavity. Normally the death of cell and the growth are active processes and depend not only on external factors but also on the expression of genes such as Bcl-2, which activate and inhibit apoptosis. The term Bcl-2 is an acronym for B-cell lymphoma/leukemia-2 genes. It has been reported that there is deregulation of Bcl-2 expression during progression from oral epithelial dysplasia to squamous cell carcinoma.Expression of this oncoprotein can be detected by immunohistochemistry. Aims and objectivesAn attempt was made to evaluate Bcl-2 oncoprotein expression in patients with oral precancer and cancer. Materials and methodsA selective prospective clinical and immunohistochemical study. Clinicopathological examination was correlated with immunohistochemical findings. The immunolocalization of Bcl-2 protein was performed using the labeled streptavidin biotin method. To visualize the reaction, 3,3-diaminobenzidine was used. ResultsBcl-2 expression was positive in 11 [36.66%, low Bcl-2 expression 3 (10.00%), moderate Bcl-2 expression 7 (23.33%), and high Bcl-2 expression 1 (3.33%)] oral cancer cases and 14 [87.50%, low expression 8 (50%), moderate expression 6 (37.50%)] precancer cases. ConclusionOn the basis of the results of our study, we conclude that positive Bcl-2 expression may be an indicator of poor prognosis in oral cancer and precancer.

High Dose Chemotherapy with Autograft in First Relapse May Overcome the Poor Prognosis of Diffuse Large B-Cell Lymphoma Patients with MYC/BCL2 Co-Expression

The immunohistochemical (ICH) co-expression of MYC and BCL2 (double expressors, DE) has been emerging as a strong and reproducible risk factor in diffuse large B cell lymphoma (DLBCL) patients (pts) treated with R-CHOP. Our aim was to analyze the prognostic value of the co-expression in DLBCL pts treated with high-dose chemotherapy followed by autologous stem cell transplantation at first relapse.In this retrospective study, we analyzed a cohort of 64 young [median age 53, (range18-70)] and/or fit DLBCL pts treated between 2003 and 2015 at the Istituto Nazionale dei Tumori Milano. All pts relapsed at median time of 4.9 months after a Rituximab-based chemotherapy (48 R-CHOP-like and 16 other intensive regimens). All pts were considered fit and eligible for high-dose chemotherapy salvage treatment. Tumor samples were analyzed by ICH for MYC and BCL2 (40%/70% threshold). Germinal B-Cell (GBC) and Activated B-Cell (ABC) DLBCL classification was done with ICH as established by the Hans algorithm. Pts characteristics at relapse were: refractory: 37/64 (58%), Ann Arbor stage≥3: 42/64 (65%), IPI score≥2: 26/60 (43%); Extranodal disease: 37/64 (58%); Bulky (defined as max diameter ≥5 cm): 29/64 (45%); CNS involvement: 8/64 (12.5%).Twenty-nine of 64 (45%) pts responded to salvage high-dose therapy (CR,PR) and 25 of them underwent autologous stem cell transplantation (ASCT). Among 39 (61%) refractory pts to first salvage intensive chemotherapy approaches, 11 (28%) responded after third or fourth chemo-immunotherapy salvage lines and underwent ASCT as well. Overall the reasons for not receiving ASCT were: refractory disease (n=20), poor mobilization (n=3), the occurrence of significant infection or toxic complications during the salvage therapy (n=4) or patient decision (n=1). ICH MYC and BCL2 positive expression was observed in 31 (48%) and 39 (61%) pts respectively. Among them, 17 (26%) were characterized by BCL2 and MYC co-expression. According with Hans algorithm, 23 (56%) and 18 (44%) DLBCL pts were classified as GBC and ABC-DLBCL. DE pts did not show any significant differences compared to others in terms of age, IPI score, presence of extranodal disease, BM infiltration, CNS involvement at relapse, refractory disease or relapse occurrence less than 1 year after first line therapy, and proportion of ABC-DLBCL subtype. Considering the whole pts cohort 5-years progression free survival (PFS) and overall survival (OS) were 27.3% (95% CI, 21.1%-33,4%) and 40.6% (95% CI, 33.2%-48%) respectively. In univariate analysis, pts with disease relapsed less than 1 year after first line therapy were characterized by a significant worse outcome in terms of 5-years PFS [11.5% (95% IC 6.1%-16.9%) vs 72.2% (95% IC 60%-84.4%] and OS [24.5% (95% IC 16.9%-32.1%) vs 84.4% (95% IC 74%-94.8%)] (p=0.0001 and p=0.0009 respectively). Similar results were observed comparing pts with refractory disease to first line therapy with others [5-years PFS 11.7% (95% IC, 5.9%-17.5%) vs 48.2% (95% IC, 37.1%-59.3%)] and 5-years OS [19.9% (95% IC, 11.8%-28%) vs 67.9% (95% IC, 57.6%-78.2%)] (p=0.0003 and p=0.001 respectively). Other variables at first relapse associated with worse outcome in terms of PFS and OS were IPI score ≥2 and presence of bulky disease. In contrast, DE patients did not shown any significant differences compared to other patents in term of 5-years PFS [39.7 (95% IC, 27.5-51.9) vs 25.1 (95% IC, 18.4-31.8), p=0.5] and 5-years OS [40.8 (95% IC, 32.3-49.3) vs 22.2 (95% IC, 5-39.4), p=0.8]. Furthermore singularly ICH MYC or BCL2 expression did not influence neither PFS nor OS.These data suggest that salvage programs with high dose chemotherapy with autograft in first relapse may overcome the previously reported poor prognosis associated with ICH MYC/BCL2 co-expression. The most significant and robust prognostic factor among DLBCL pts in first relapse is still represented by refractoriness and the time of relapse. DisclosuresNo relevant conflicts of interest to declare.

Expression of c-erb-B2 gene in bladder cancer of Egyptian patients and its correlation with p53 and bcl-2.

Urinary bladder cancer is the 9th most common type of cancer and the 13th most common cause of death worldwide. C-erbB-4 is a class of oncogenes plays a role in cancer development. The present work was performed to assess C-erbB-4 oncogene amplification by PCR technology and its correlation with p53 and bcl-2. This study included 50 male patients (10 controls and 40 urinary bladder cancer patients). The bladder cancer patients include 20 specimens diagnosed as transitional cell carcinoma (TCC) and 20 specimens diagnosed as squamous cell carcinoma (SCC). The results revealed that 7 (35%) of both TCC and SCC showed c-erb-B2 gene amplification. 12 (60%) of TCC and 6 (30%) of SCC showed positive expression of p53. 11 (55%) of TCC and 6 (30%) of SCC showed positive Bcl-2 expression. A direct statistically significant association was detected between c-erb-B2 expression and Bcl-2 and p53 expression in TCC and SCC specimens. Seven (35%) of TCC showed c-erb-B2 gene amplification and expression of both p53 and Bcl-2. Five (25%) of the examined SCC specimens showed c-erb-B2 gene amplification and positive expression for both p53 and Bcl-2. The results indicated that a direct statistically significant association was detected in TCC group between amplification of c-erb-B2 gene by PCR and expressionof p53 and Bcl-2 by immunohistochemistry.