Lactate-induced up-regulation of PLEKHA4 promotes proliferation and apoptosis of human glioma cells / 南方医科大学学报

OBJECTIVE@#To investigate the effect of lactic acid-induced upregulation of PLEKHA4 expression on biological behaviors of glioma cells and the possible molecular mechanism.@*METHODS@#GEO database and GEPIA2 website were used to analyze the relationship between PLEKHA4 expression level and the pathological grade of glioma. A specific PLEKHA4 siRNA was transfected in glioma U251 and T98G cells, and the changes in cell proliferation ability were assessed by real-time cell analysis technology and Edu experiment. The colony-forming ability of the cells was evaluated using plate cloning assay, and cell cycle changes and cell apoptosis were analyzed with flow cytometry. The mRNA expression of PLEKHA4 was detected by PCR in glioma samples and controls and in glioma cells treated with lactic acid and glucose. Xenograft mice in vivo was used to detect tumor formation in nude mice; Western blotting was used to detect the expressions of cyclinD1, CDK2, Bcl2, β-catenin and phosphorylation of the key proteins in the MAPK signaling pathway.@*RESULTS@#The results of GEO database and online website analysis showed that PLEKHA4 was highly expressed in glioma tissues and was associated with poor prognosis; PLEKHA4 knockdown obviously inhibited the proliferation and attenuated the clone-forming ability of the glioma cells (P < 0.05). Flow cytometry showed that PLEKHA4 knockdown caused cell cycle arrest in G1 phase and promoted apoptosis of the cells (P < 0.01). PLEKHA4 gene mRNA expression was increased in glioma samples and glioma cells after lactate and glucose treatment (P < 0.01). PLEKHA4 knockdown, tumor formation ability of nude mice decreased; PLEKHA4 knockdown obviously lowered the expression of cyclinD1, CDK2, Bcl2 and other functional proteins, inhibited the phosphorylation of ERK and p38 and reduced the expression of β-catenin protein (P < 0.01).@*CONCLUSION@#PLEKHA4 knockdown inhibited the proliferation of glioma cells and promoted apoptosis by inhibiting the activation of the MAPK signaling pathway and expression of β-catenin. Lactic acid produced by glycolysis upregulates the expression of PLEKHA4 in glioma cells.

Therapeutic mechanism of Guizhi Gancao Decoction for heart failure: a network pharmacology-based analysis / 南方医科大学学报

OBJECTIVE@#To predict the targets and pathways in the therapeutic mechanism of Guizhi Gancao Decoction (GZGCD) against heart failure (HF) based on network pharmacology.@*METHODS@#The chemical components of GZGCD were analyzed using the databases including TCMSP, TCMID and TCM@Taiwan, and the potential targets of GZGCD were predicted using the SwissTargetPrediction database. The targets of HF were obtained using the databases including DisGeNET, Drugbank and TTD. The intersection targets of GZGCD and HF were identified using VENNY. Uniport database was used to convert the information, and the components-targets-disease network was constructed using Cytoscape software. The Bisogene plug-in, Merge plug-in, and CytoNCA plug-in in Cytoscape software were used for protein-protein interaction (PPI) analysis to acquire the core targets. Metascape database was used for GO and KEGG analysis. The results of network pharmacology analysis were verified with Western blot analysis. Three factors (PKCα, ERK1/2 and BCL2) were screened according to the degree value of network pharmacology results and the degree of correlation with heart failure process. The pentobarbtal sodium was dissolvein H9C2 cells treated with serum-free high glucose medium to simulate the ischemic anoxic environment of heart failure. The total proteins of myocardial cells were extracted. The protein contents of PKCα, ERK1/2 and BCL2 were determined.@*RESULTS@#We identified a total of 190 intersection targets between GZGCD and HF using Venny database, involving mainly the circulatory system process, cellular response to nitrogen compounds, cation homeostasis, and regulation of the MAPK cascade. These potential targets were also involved in 38 pathways, including the regulatory pathways in cancer, calcium signal pathway, cGMP-PKG signal pathway, and cAMP signal pathway. Western blot analysis showed that in an in vitro H9C2 cell model of HF, treatment with GZGCD downregulated PKCα and ERK1/2 expressions and upregulated BCL2 expression.@*CONCLUSION@#The therapeutic mechanism of GZGCD for HF involves multiple targets including PRKCA, PRKCB, MAPK1, MAPK3, and MAPK8 and multiple pathways including the regulatory pathway in cancer and the calcium signaling pathway.

Clinicopathological features of fibrin-associated diffuse large B-cell lymphoma: a report of six cases / 中华病理学杂志

Objective: To investigate the clinical, pathological and immunophenotypic features, molecular biology and prognosis of fibrin-associated large B-cell lymphoma (LBCL-FA) in various sites. Methods: Six cases of LBCL-FA diagnosed from April 2016 to November 2021 at the Beijing Friendship Hospital, Capital Medical University, Beijing, China and the First Affiliated Hospital, Wenzhou Medical University, Wenzhou, China were collected. The cases were divided into atrial myxoma and cyst-related groups. Clinical characteristics, pathological morphology, immunophenotype, Epstein Barr virus infection status, B-cell gene rearrangement and fluorescence in situ hybridization of MYC, bcl-2, bcl-6 were summarized. Results: The patients' mean age was 60 years. All of them were male. Three cases occurred in atrial myxoma background, while the others were in cyst-related background, including adrenal gland, abdominal cavity and subdura. All cases showed tumor cells located in pink fibrin clot. However, three cyst-related cases showed the cyst wall with obviously fibrosis and inflammatory cells. All cases tested were non germinal center B cell origin, positive for PD-L1, EBER and EBNA2, and were negative for MYC, bcl-2 and bcl-6 rearrangements, except one case with MYC, bcl-2 and bcl-6 amplification. All of the 5 cases showed monoclonal rearrangement of the Ig gene using PCR based analysis. The patients had detailed follow-ups of 9-120 months, were treated surgically without radiotherapy or chemotherapy, and had long-term disease-free survivals. Conclusions: LBCL-FA is a group of rare diseases occurring in various sites, with predilection in the context of atrial myxoma and cyst-related lesions. Cyst-related lesions with obvious chronic inflammatory background show more scarcity of lymphoid cells and obvious degeneration, which are easy to be missed or misdiagnosed. LBCL-FA overall has a good prognosis with the potential for cure by surgery alone and postoperative chemotherapy may not be necessary.

Alcohol extract of root and root bark of Toddalia asiatica alleviates CIA in rats through anti-inflammatory and proapoptotic effects / 中国中药杂志

This study aims to investigate the therapeutic effect of alcohol extract of root and root bark of Toddalia asiatica(TAAE) on collagen-induced arthritis(CIA) in rats through phosphatidylinoinosidine-3 kinase/protein kinase B(PI3K/Akt) signaling pathway. To be specific, CIA was induced in rats, and then the rats were treated(oral, daily) with TAAE and Tripterygium Glycoside Tablets(TGT), respectively. The swelling degree of the hind leg joints was scored weekly. After 35 days of administration, the histopathological changes were observed based on hematoxylin and eosin(HE) staining. Enzyme-linked immunosorbent assay(ELISA) was employed to detect the levels of cytokines [tumor necrosis factor-α(TNF-α), interleukin(IL)-6)]. Terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL) staining was performed to detect the apoptosis of synoviocytes in rats. Western blot was used to detect the expression levels of apoptosis-related proteins B-cell lymphoma 2(Bcl-2)-associated X(Bax), Bcl-2, and caspase-3 and pathway-related proteins phosphoinositide 3-kinase(PI3K), phosphorylated(p)-PI3K, protein kinase B(Akt), and p-Akt. RT-qPCR was conducted to examine the mRNA levels of Bax, Bcl-2, caspase-3, TNF-α, IL-6, and IL-1β and pathway-related proteins PI3K, p-PI3K, Akt, and p-Akt. TAAE can alleviate the joint swelling in CIA rats, reduce serum levels of inflammatory cytokines, improve synovial histopathological changes, promote apoptosis of synoviocytes, and inhibit synovial inflammation. In addition, RT-qPCR and Western blot results showed that TAAE up-regulated the level of Bax, down-regulated the level of Bcl-2, and activated caspase-3 to promote apoptosis in synoviocytes. TAAE effectively down-regulated the protein levels of p-PI3K and p-Akt. In this study, TAAE shows therapeutic effect on CIA in rats and reduces the inflammation. The mechanism is that it suppresses PI3K/Akt signaling pathway and promotes synoviocyte apoptosis. Overall, this study provides a new clue for the research on the anti-inflammatory mechanism of TAAE and lays a theoretical basis for the better clinical application of TAAE in the treatment of inflammatory and autoimmune diseases.

Stellera chamaejasme extract against multidrug resistance of breast cancer cell line MCF-7 / 中国中药杂志

This study explored the effect and underlying mechanism of Stellera chamaejasme extract(SCE) on multidrug resistance of breast cancer. The chemotherapy-sensitive breast cancer cell line MCF-7 and adriamycin(ADR)-resistant cell line MCF-7/ADR were used as experimental subjects. MTT assay was used to detect cell proliferation activity. Pi staining was used to detect the cell cycle. 4',6-Diamidino-2-phenylindole, dihydrochloride(DAPI) staining and flow cytometry were used to detect apoptosis. Dansylcadaverine(MDC) staining and GFP-LC3B-Mcherry adenovirus transfection were used to detect autophagy. The protein expression of Bcl-2, Bax, caspase-9, caspase-3, LC3B, p62, and Beclin-1 was detected by Western blot. The results showed that SCE could significantly inhibit the proliferation of both sensitive and resistant breast cancer cell lines. The drug resistance factor was 0.53, which was significantly lower than 59 of ADR. Meanwhile, the proportion of sensitive/resistant cells in the G_0/G_1 phase increased significantly after SCE treatment. In addition, DAPI staining showed that a series of apoptosis phenomena such as nuclear pyknosis, staining deepening, and nuclear fragmentation appeared in sensitive/resistant cell lines after SCE administration. Moreover, the results of flow cytometry double staining showed that the proportion of apoptotic cells in sensitive/resistant cell lines increased significantly after SCE administration. Besides, Western blot showed that the protein expression levels of caspase-3, caspase-9, and Bcl-2 significantly decreased and the expression level of Bax protein significantly increased in both breast cancer cell lines after SCE administration. Furthermore, SCE could also increase the positive fluorescent spots after MDC staining and yellow fluorescent spots after GFP-LC3B-mcherry transfection, and up-regulate the expression levels of autophagy-related proteins LC3B-Ⅱ, p62, and Beclin-1 in breast cancer cells. In summary, SCE may play the role of anti-multidrug resistance by blocking the cell cycle of breast cancer multidrug-resistant cells, blocking autophagy flow, and ultimately interfering with the apoptosis resistance of drug-resistant cells.

Acteoside promotes autophagy and apoptosis of hepatoma cells by regulating JNK signaling pathway / 中国中药杂志

This study explored the molecular mechanism of acteoside against hepatoma 22(H22) tumor in mice through c-Jun N-terminal kinase(JNK) signaling pathway. H22 cells were subcutaneously inoculated in 50 male BALB/c mice, and then the model mice were classified into model group, low-dose, medium-dose, and high-dose acteoside groups, and cisplatin group. The administration lasted 2 weeks for each group(5 consecutive days/week). The general conditions of mice in each group, such as mental status, diet intake, water intake, activity, and fur were observed. The body weight, tumor volume, tumor weight, and tumor-inhibiting rate were compared before and after administration. Morphological changes of liver cancer tissues were observed based on hematoxylin and eosin(HE) staining, and the expression of phosphorylated(p)-JNK, JNK, B-cell lymphoma-2(Bcl-2), Beclin-1, and light chain 3(LC3) in each tissue was detected by immunohistochemistry and Western blot. qRT-PCR was performed to detect the mRNA expression of JNK, Bcl-2, Beclin-1, and LC3. The general conditions of mice in model and low-dose acteoside groups were poor, while the general conditions of mice in the remaining three groups were improved. The body weight of mice in medium-dose acteoside group, high-dose acteoside group, and cisplatin group was smaller than that in model group(P<0.01). The tumor volume in model group was insignificantly different from that in low-dose acteoside group, and the volume in cisplatin group showed no significant difference from that in high-dose acteoside group. Tumor volume and weight in medium-dose and high-dose acteoside groups and cisplatin group were lower than those in the model group(P<0.001). The tumor-inhibiting rates were 10.72%, 40.32%, 53.79%, and 56.44% in the low-dose, medium-dose, and high-dose acteoside groups and cisplatin group, respectively. HE staining showed gradual decrease in the count of hepatoma cells and increasing sign of cell necrosis in the acteoside and cisplatin groups, and the necrosis was particularly obvious in the high-dose acteoside group and cisplatin group. Immunohistochemical results suggested that the expression of Beclin-1, LC3, p-JNK, and JNK was up-regulated in acteoside and cisplatin groups(P<0.05). The results of immunohistochemistry, Western blot, and qRT-PCR indicated that the expression of Bcl-2 was down-regulated in the medium-dose and high-dose acteoside groups and cisplatin group(P<0.01). Western blot showed that the expression of Beclin-1, LC3, and p-JNK was up-regulated in acteoside and cisplatin groups(P<0.01), and there was no difference in the expression of JNK among groups. qRT-PCR results showed that the levels of Beclin-1 and LC3 mRNA were up-regulated in the acteoside and cisplatin groups(P<0.05), and the level of JNK mRNA was up-regulated in medium-dose and high-dose acteoside groups and cisplatin group(P<0.001). Acteoside promotes apoptosis and autophagy of H22 cells in mice hepatoma cells by up-regulating the JNK signaling pathway, thus inhibiting tumor growth.

Tanshinone IIA inhibits hypoxia/reoxygenation-induced cardiomyocyte apoptosis and autophagy by regulating ABCE1 / 中华危重病急救医学

OBJECTIVE@#To investigate the effects of tanshinone IIA on apoptosis and autophagy induced by hypoxia/reoxygenation in H9C2 cardiomyocytes and its mechanism.@*METHODS@#H9C2 cardiomyocytes in logarithmic growth phase were divided into control group, hypoxia/reoxygenation model group and tanshinone IIA low-dose, medium-dose and high-dose groups (50, 100, 200 mg/L tanshinone IIA were treated after hypoxia/reoxygenation respectively). The dose with good therapeutic effect was selected for follow-up study. The cells were divided into control group, hypoxia/reoxygenation model group, tanshinone IIA+pcDNA3.1-NC group and tanshinone IIA+pcDNA3.1-ABCE1 group. The cells were transfected with the overexpressed plasmids pcDNA3.1-ABCE1 and pcDNA3.1-NC and then treated accordingly. Cell counting kit-8 (CCK-8) was used to detect H9C2 cell activity in each group. The apoptosis rate of cardiomyocytes was detected by flow cytometry. The ATP-binding cassette transporter E1 (ABCE1), apoptosis-related proteins Bcl-2 and Bax, caspase-3, autophagy-related proteins Beclin-1, microtubule-associated protein 1 light chain 3 (LC3II/I) and p62 mRNA expression level of H9C2 cells in each group were detected by real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR). The protein expression levels of the above indexes in H9C2 cells were detected by Western blotting.@*RESULTS@#(1) Cell activity and ABCE1 expression: tanshinone IIA inhibited the activity of H9C2 cells induced by hypoxia/reoxygenation, and the effect was significant at medium-dose [(0.95±0.05)% vs. (0.37±0.10)%, P < 0.01], mRNA and protein expression of ABCE1 were significantly reduced [ABCE1 mRNA (2-ΔΔCt): 2.02±0.13 vs. 3.74±0.17, ABCE1 protein (ABCE1/GAPDH): 0.46±0.04 vs. 0.68±0.07, both P < 0.05]. (2) Expression of apoptosis-related proteins: medium-dose of tanshinone IIA inhibited the apoptosis of H9C2 cells induced by hypoxia/reoxygenation [apoptosis rate: (28.26±2.52)% vs. (45.27±3.07)%, P < 0.05]. Compared with the hypoxia/reoxygenation model group, medium-dose of tanshinone IIA significantly down-regulated the protein expression of Bax and caspase-3 in H9C2 cells induced by hypoxia/reoxygenation, and significantly up-regulated the protein expression of Bcl-2 [Bax (Bax/GAPDH): 0.28±0.03 vs. 0.47±0.03, caspase-3 (caspase-3/GAPDH): 0.31±0.02 vs. 0.44±0.03, Bcl-2 (Bcl-2/GAPDH): 0.53±0.02 vs. 0.37±0.05, all P < 0.05]. (3) Expression of autophagy-related proteins: compared with the control group, the positive rate of LC3 in the hypoxia/reoxygenation model group was significantly increased, while the positive rate of LC3 in the medium-dose of tanshinone IIA group was significantly decreased [(20.67±3.09)% vs. (42.67±3.86)%, P < 0.01]. Compared with hypoxia/reoxygenation model group, medium-dose of tanshinone IIA significantly down-regulated Beclin-1, LC3II/I and p62 protein expressions [Beclin-1 (Beclin-1/GAPDH): 0.27±0.05 vs. 0.47±0.03, LC3II/I ratio: 0.24±0.05 vs. 0.47±0.04, p62 (p62/GAPDH): 0.21±0.03 vs. 0.48±0.02, all P < 0.05]. (4) Expression of apoptosis and autophagy related proteins after transfection with overexpressed ABCE1 plasmid: compared with tanshinone IIA+pcDNA3.1-NC group, the protein expression levels of Bax, caspase-3, Beclin-1, LC3II/I and p62 in tanshinone IIA+pcDNA3.1-ABCE1 group were significantly up-regulated, while the protein expression level of Bcl-2 was significantly down-regulated.@*CONCLUSIONS@#100 mg/L tanshinone IIA could inhibit autophagy and apoptosis of cardiomyocytes by regulating the expression level of ABCE1. So, it protects H9C2 cardiomyocytes injury induced by hypoxia/reoxygenation.

Experimental Study on the Mechanism of Mangiferin Inhibiting Malignant Biological Characteristics of Multiple Myeloma and Exerting Anticancer Effect / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of pure Chinese herbal extract Mangiferin on the malignant biological behaviors of multiple myeloma (MM) cells, and to analyze the molecular mechanism of the anti-myeloma effect of Mangiferin, so as to provide experimental basis for MM replacement therapy.@*METHODS@#U266 and RPMI8226 of human MM cell lines were intervened with different concentrations of Mangiferin. Cell proliferation was detected by CCK-8 method. Annexin V/PI double staining flow cytometry was used to detect cell apoptosis. Western blot was used to detect the expression of apoptosis and related signaling pathway proteins, and real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of matrix metalloproteinase (MMP) and CXC chemokine receptor (CXCR) family.@*RESULTS@#Mangiferin could inhibit the proliferation activity of U266 and RPMI8226 cells and induce cells apoptosis. After Mangiferin intervened in U266, RPMI8226 cells for 48 h, the expression of Bcl-2 family pro-apoptotic protein Bax was up-regulated, while the expression of survivin and Bcl-xL proteins was down-regulated and caspase-3 was hydrolyzed and activated to promote cell apoptosis, besides, the expression of Bcl-2 protein in U266 cells was also significantly down-regulated to induce apoptosis (P<0.05). After Mangiferin intervenes in MM cells, it can not only increase the expression level of tumor suppressor p53, but also induce programmed cell death of MM cells by inhibiting the expression of anti-apoptotic molecules and down-regulating the phosphorylation levels of AKT and NF-κB. In addition, after the intervention of Mangiferin, the expressions of CXCR4, MMP2 and MMP9 in U266 cells were down-regulated (P<0.05), while there is no effect on the expressions of CXCR2, CXCR7 and MMP13 (P>0.05). However, the expressions of CXCR4, MMP9, and MMP13 in RPMI8226 cells were down-regulated (P<0.01), the expression of MMP2 was weakly affected, and the expression of CXCR2 and CXCR7 was basically not affected (P>0.05).@*CONCLUSION@#Mangiferin can inhibit the proliferation and induce apoptosis of MM cells, and its mechanism may be related to inhibiting the activation of NF-κB signaling pathway, affecting the expression of Bcl-2 family proteins, and inhibiting the expression of core members of MMP and CXCR family.

Establishment and Clinical Significance of Prognostic Nomogram Model for Diffuse Large B-Cell Lymphoma Based on Immunohistochemistry Markers and International Prognostic Index Scores / 中国实验血液学杂志

OBJECTIVE@#To retrospectively analyze clinical characteristics and survival time of patients with diffuse large B-cell lymphoma (DLBCL), detect prognosis-related markers, and establish a nomogram prognostic model of clinical factors combined with biomarkers.@*METHODS@#One hundred and thirty-seven patients with DLBCL were included in this study from January 2014 to March 2019 in the First Affiliated Hospital of Nanchang University. The expression of GCET1, LMO2, BCL-6, BCL-2 and MYC protein were detected by immunohistochemistry (IHC), then the influences of these proteins on the survival and prognosis of the patients were analyzed. Univariate and multivariate Cox regression analysis were used to gradually screen the prognostic factors in nomogram model. Finally, nomogram model was established according to the result of multivariate analysis.@*RESULTS@#The positive expression of GCET1 protein was more common in patients with Ann Arbor staging I/II (P =0.011). Compared with negative patients, patients with positive expression of LMO2 protein did not often show B symptoms (P =0.042), and could achieve better short-term curative effect (P =0.005). The overall survival (OS) time of patients with positive expression of LMO2 protein was significantly longer than those with negative expression of LMO2 protein (P =0.018), though the expression of LMO2 protein did not correlate with progression-free survival (PFS) (P >0.05). However, the expression of GCET1 protein had no significant correlation with OS and PFS. Multivariate Cox regression analysis showed that nomogram model consisted of 5 prognostic factors, including international prognostic index (IPI), LMO2 protein, BCL-2 protein, MYC protein and rituximab. The C-index applied to the nomogram model for predicting 4-year OS rate was 0.847. Moreover, the calibrated curve of 4-year OS showed that nomogram prediction had good agreement with actual prognosis.@*CONCLUSION@#The nomogram model incorporating clinical characteristics and IHC biomarkers has good discrimination and calibration, which provides a useful tool for the risk stratification of DLBCL.

Regulation of Baicalin on Growth of Extranodal NK/T Cell Lymphoma Cells through FOXO3/CCL22 Signaling Pathway / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of baicalin on the growth of extranodal NK/T cell lymphoma (ENKTCL) cells and its related mechanism.@*METHODS@#Normal NK cells and human ENKTCL cells lines SNK-6 and YTS were cultured, then SNK-6 and YTS cells were treated with 5, 10, 20 μmol/L baicalin and set control. Cell proliferation and apoptosis was detected by Edu method and FCM method, respectively, and expressions of BCL-2, Bax, FOXO3 and CCL22 proteins were detected by Western blot. Interference plasmids were designed and synthesized. FOXO3 siRNA interference plasmids and CCL22 pcDNA overexpression plasmids were transfected with PEI transfection reagent. Furthermore, animal models were established for validation.@*RESULTS@#In control group and 5, 10, 20 μmol/L baicalin group, the proliferation rate of SNK-6 cells was (56.17±2.96)%, (51.92±4.63)%, (36.42±1.58)%, and (14.60±2.81)%, respectively, while that of YTS cells was (58.85±2.98)%, (51.38±1.32)%, (34.75±1.09)%, and (15.45±1.10)%, respectively. In control group and 5, 10, 20 μmol/L baicalin group, the apoptosis rate of SNK-6 cells was (5.93±0.74)%, (11.78±0.34)%, (28.46±0.44)%, and (32.40±0.37)%, respectively, while that of YTS cells was (7.93±0.69)%, (16.29±1.35)%, (33.91±1.56)%, and (36.27±1.06)%, respectively. Compared with control group, the expression of BCL-2 protein both in SNK-6 and YTS cells decreased significantly (P<0.001), and the expression of Bax protein increased in SNK-6 cells only when the concentration of baicalin was 20 μmol/L (P<0.001), while that in YTS cells increased in all three concentrations(5, 10, 20 μmol/L) of baicalin (P<0.001). The expression of FOXO3 protein decreased while CCL22 protein increased in ENKTCL cell lines compared with human NK cells (P<0.001), but the expression of FOXO3 protein increased (P<0.01) and CCL22 protein decreased after baicalin treatment (P<0.001). Animal experiments showed that baicalin treatment could inhibit tumor growth. The expression of CCL22 protein in ENKTCL tissue of nude mice treated with baicalin decreased compared with control group (P<0.01), while the FOXO3 protein increased (P<0.05). In addition, FOXO3 silencing resulted in the decrease of FOXO3 protein expression and increase of CCL22 protein expression (P<0.01, P<0.001).@*CONCLUSION@#Baicalin can inhibit proliferation and promote apoptosis of ENKTCL cell lines SNK-6 and YTS, up-regulate the expression of Bax protein, down-regulate the expression of BCL-2 protein, and down-regulate the expression of CCL22 protein mediated by FOXO3. Animal experiment shown that the baicalin can inhibit tumor growth. Baicalin can inhibit the growth and induce apoptosis of ENKTCL cells through FOXO3/CCL22 signaling pathway.

The Effects and Regulatory Mechanism of Targeting CXC Chemokine Receptor 1/2 Combined with Ara-C on the Malignant Biological Behaviors of U937 Cells of Acute Myeloid Leukemia / 中国实验血液学杂志

OBJECTIVE@#To investigate and analyze the effect of CXC chemokine receptor 1/2 (CXCR1/2) targeting inhibitor Reparixin combined with cytarabine (Ara-C) on the malignant biological behaviors of acute myeloid leukemia cells and its effect on the expression of the CXCR family, while exploring the accompanying molecular mechanism, providing scientific basis and reference for new molecular markers and targeted therapy for AML.@*METHODS@#Acute myeloid leukemia U937 cells were treated with different concentrations of Reparixin, Ara-C alone or in combination, and the cell morphology was observed under an inverted microscope; Wright-Giemsa staining was used to detect cell morphological changes; CCK-8 method was used to detect cell proliferation; the ability of cell invasion was detected by Transwell chamber method; the ability of colony formation was detected by colony formation assay; cell apoptosis was detected by Hoechst 33258 fluorescent staining and Annexin V/PI double-staining flow cytometry; monodansylcadaverine(MDC) staining was used to detect cell autophagy; the expression of apoptosis, autophagy and related signaling pathway proteins was detected by Western blot and the expression changes of CXCR family were detected by real-time quantitative polymerase chain reaction (qRT-PCR).@*RESULTS@#Reparixin could inhibit the proliferation, invasion, migration and clone formation ability of U937 cells. Compared with the single drug group, when U937 cells were intervened by Reparixin combined with Ara-C, the malignant biological behaviors such as proliferation, invasion and colony formation were significantly decreased, and the levels of apoptosis and autophagy were significantly increased (P<0.01). After Reparixin combined with Ara-C intervenes in U937 cells, it can up-regulate the expression of the pro-apoptotic protein Bax and significantly down-regulate the expression of the anti-apoptotic protein Bcl-2, and also hydrolyze and activate Caspase-3, thereby inducing cell apoptosis. Reparixin combined with Ara-C could up-regulate the expressions of LC3Ⅱ and Beclin-1 proteins in U937 cells, and the ratio of LC3Ⅱ/LC3Ⅰ in cells was significantly up-regulated compared with single drug or control group (P<0.01). MDC result showed that the green granules of vesicles increased significantly, and a large number of broken cells were seen (P<0.01). Reparixin combined with Ara-C can significantly inhibit the phosphorylation level of PI3K, AKT and NF-κB signaling molecule, inhibit the malignant biological behavior of cells by inhibiting the activation of PI3K/AKT/NF-κB pathway, and induce programmed cell death. Ara-C intervention in U937 cells had no effect on the expression of CXCR family (P>0.05). The expression of CXCR1, CXCR2, and CXCR4 mRNA could be down-regulated by Reparixin single-agent intervention in U937 cells (P<0.05), and the expression of CXCR2 was more significantly down-regulated than the control group and other CXCRs (P<0.01). When Reparixin and Ara-C intervened in combination, the down-regulated levels of CXCR1 and CXCR2 were more significant than those in the single-drug group (P<0.01), while the relative expressions of CXCR4 and CXCR7 mRNA had no significant difference compared with the single-drug group (P>0.05).@*CONCLUSION@#Reparixin combined with Ara-C can synergistically inhibit the malignant biological behaviors of U937 cells such as proliferation, invasion, migration and clone formation, and induce autophagy and apoptosis. The mechanism may be related to affecting the proteins expression of Bcl-2 family and down-regulating the proteins expression of CXCR family, while inhibiting the PI3K/AKT/NF-κB signaling pathway.

Anti-glioma mechanism of pterostilbene by regulating apoptosis and GSDME-mediated pyroptosis pathways: a study based on network pharmacology and experimental research / 中国中药杂志

This study aimed to explore the anti-glioma effect of natural compound pterostilbene(PTE) through regulating pyroptosis and apoptosis pathways, and to analyze the possible anti-glioma pathways and targets of PTE by network pharmacology and molecular docking. In this study, the action targets of PTE and the glioma targets were obtained by network pharmacology to construct a target network and a protein-protein interaction(PPI) network to predict the possible action targets of PTE against glioma. Molecular docking was performed on the core targets by AutoDock and the action pathways of PTE against glioma were predicted by enrichment analysis. In addition, the effect of PTE on the viability of U87MG and GL261 glioma cells was detected by CCK-8 assay. Clone formation assay and cell scratching assay were used to explore the effect of different concentrations of PTE on the proliferation and migration, respectively of glioma cells. Hoechst staining was used to observe PTE-induced apoptosis in glioma cells. The changes in mitochondrial membrane potential were detected by JC-1 staining. The pyroptosis-inducing effect of PTE on glioma cells was observed by inverted microscopy and lactate dehydrogenase(LDH) assay. Hoechst 33342/PI dual staining assay was performed to detect the integrity of glioma cell membranes. The expressions of pyroptosis and apoptosis-related proteins in glioma cells after PTE induction were determined by Western blot. In this study, 37 anti-glioma targets of PTE were obtained, and enrichment analysis suggested that PTE exerted anti-glioma effects through various signaling pathways including cancer pathway, proteoglycan in cancer, PI3K/AKT pathway, and apoptosis regulatory pathway. Molecular docking revealed that PTE had good binding activity with the main targets. Compared with the control group, PTE significantly reduced the viability as well as the proliferation, migration and adhesion abilities of U87MG and GL261 cells; it induced the apoptosis of the two glioma cells and the decrease of mitochondrial membrane potential in U87MG cells, and the effects increased with the increase of drug concentration. Compared with the conditions in the control group, glioma cells in the PTE group had increased pyroptosis-specific appearance and gradually increased LDH release; the number of PI positive cells was significantly elevated with the increase of PTE concentration as revealed by Hoechst 33342/PI staining; the expression levels of apoptosis-related factors cleaved PARP1 and B-cell lymphoma-2(Bcl-2) associated X(BAX) in the PTE group were markedly up-regulated, while the expression level of Bcl-2 was markedly down-regulated; the activation levels of pyroptosis-related proteins cleaved caspase-3 and gasdermin E-N(GSDME-N) had a remarkable rise in the PTE group, while no significant changes were found in the activation levels of gasdermin D-N(GSDMD-N) and cleaved caspase-1. In summary, PTE plays an anti-glioma role by inhibiting cell viability, proliferation, and migration and activating the caspase-3/GSDME-mediated pyroptosis pathway and mitochondrial apoptosis pathway.

Effect and mechanism of Xihuang Pills on rats with precancerous lesions of breast / 中国中药杂志

The purpose of this study was to explore the effect and mechanism of Xihuang Pills on rats with precancerous lesions of the breast. Of 48 healthy female rats, 8 were randomly selected as blank group, and the other 40 were treated with 7,12-dimethylbenzanthracene(DMBA) combined with estrogen and progestin to establish a model of precancerous lesions of the breast. The successfully modeled rats were randomly divided into a model group, a tamoxifen group(1.8 mg·kg~(-1)·d~(-1)), a Xihuang Pills low-dose group(0.3 g·kg~(-1)·d~(-1)), a medium-dose group(0.6 g·kg~(-1)·d~(-1)) and a high-dose group(1.2 g·kg~(-1)·d~(-1)). After 30 days of admi-nistration, the histopathological changes of viscera and breast were observed by haematoxylin and eosin(HE) staining, and the visceral index was calculated. Enzyme linked immunosorbent assay(ELISA) was used to detect the contents of estradiol(E_2) and progesterone(P) in serum. The protein expressions of vascular endothelial growth factor(VEGF) and fibroblast growth factor 2(FGF2) were detected by immunohistochemistry. The protein expressions of VEGF, vascular endothelial growth factor receptor 2(VEGFR2), phosphorylated-vascular endothelial growth factor receptor 2(p-VEGFR2), B-cell lymphoma-2(Bcl-2), and Bcl-2 associated X protein(Bax) were detected by Western blot and the mRNA expressions of VEGF, FGF2, CXC-chemokine receptor 4(CXCR4), cysteine aspartic acid-specific protease(caspase-3), and stromal cell-derived factor 1(SDF-1) were detected by real-time polymerase chain reaction(RT-PCR). HE staining revealed that the model group had some liver and kidney damages and severe hyperplastic mammary tissue, while the Xihuang Pills high-dose group had mild hyperplasia. Compared with the model group, the Xihuang Pills groups had lo-wer ovarian coefficient(P<0.05 or P<0.01) and Xihuang Pills high-dose group had lower uterine coefficient(P<0.01). ELISA results showed that compared with the model group, expressions of E_2 and P in Xihuang Pills high-dose group were significantly decreased(P<0.05 or P<0.01). Immunohistochemistry, Western blot and RT-PCR indicated that compared with the conditions in the model group, the protein and mRNA expressions of VEGF and FGF2 in the Xihuang Pills groups were down-regulated(P<0.05 or P<0.01), and the protein expression of Bcl-2 was lowered(P<0.01); there was a decrease in the protein expressions of VEGFR2 and p-VEGFR2(P<0.01), a down-regulation in the mRNA expressions of CXCR4 and SDF-1(P<0.01), while an increase in the mRNA expression of caspase-3(P<0.01) in both Xihuang Pills medium-dose and high-dose groups; the protein expression of Bax in Xihuang Pills high-dose group was increased(P<0.01). The above results indicated that Xihuang Pills can effectively intervene in precance-rous lesions of the breast, and the mechanism may be related to the regulation of E_2 and P secretion as well as the inhibition of angiogenesis and chemokine receptor expression, thus controlling the occurrence of precancerous lesions of the breast in rats.

Neuroprotective effect of ginsenoside Re on drosophila model of Parkinson's disease / 中国中药杂志

This study aims to explore the neuroprotective mechanism of ginsenoside Re(GS-Re) on drosophila model of Parkinson's disease(PD) induced by rotenone(Rot). To be specific, Rot was used to induce PD in drosophilas. Then the drosophilas were grouped and respectively treated(GS-Re: 0.1, 0.4, 1.6 mmol·L~(-1); L-dopa: 80 μmol·L~(-1)). Life span and crawling ability of drosophilas were determined. The brain antioxidant activity [content of catalase(CAT), malondialdehyde(MDA), reactive oxygen species(ROS), superoxide dismutase(SOD)], dopamine(DA) content, and mitochondrial function [content of adenosine triphosphate(ATP), NADH:ubiquinone oxidoreductase subunit B8(NDUFB8) Ⅰ activity, succinate dehydrogenase complex, subunit B(SDHB) Ⅱ activity] were detected by enzyme-linked immunosorbent assay(ELISA). The number of DA neurons in the brains of drosophilas was measured with the immunofluorescence method. The levels of NDUFB8 Ⅰ, SDHB Ⅱ, cytochrome C(Cyt C), nuclear factor-E2-related factor 2(Nrf2), heme oxygenase-1(HO-1), B-cell lymphoma/leukemia 2(Bcl-2)/Bcl-2-assaciated X protein(Bax), and cleaved caspase-3/caspase-3 in the brain were detected by Western blot. The results showed that model group [475 μmol·L~(-1) Rot(IC_(50))] demonstrated significantly low survival rate, obvious dyskinesia, small number of neurons and low DA content in the brain, high ROS level and MDA content, low content of SOD and CAT, significantly low ATP content, NDUFB8 Ⅰ activity, and SDHB Ⅱ activity, significantly low expression of NDUFB8 Ⅰ, SDHB Ⅱ, and Bcl-2/Bax, large amount of Cyt C released from mitochondria to cytoplasm, low nuclear transfer of Nrf2, and significantly high expression of cleaved caspase-3/caspase-3 compared with the control group. GS-Re(0.1, 0.4, and 1.6 mmol·L~(-1)) significantly improved the survival rate of PD drosophilas, alleviated the dyskinesia, increased DA content, reduced the loss of DA neurons, ROS level, and MDA content in brain, improved content of SOD and CAT and antioxidant activity in brain, maintained mitochondrial homeostasis(significantly increased ATP content and activity of NDUFB8 Ⅰ and SDHB Ⅱ, significantly up-regulated expression of NDUFB8 Ⅰ, SDHB Ⅱ, and Bcl-2/Bax), significantly reduced the expression of Cyt C, increased the nuclear transfer of Nrf2, and down-regulated the expression of cleaved caspase-3/caspase-3. In conclusion, GS-Re can significantly relieve the Rot-induced cerebral neurotoxicity in drosophilas. The mechanism may be that GS-Re activates Keap1-Nrf2-ARE signaling pathway by maintaining mitochondrial homeostasis, improves antioxidant capacity of brain neurons, then inhibits mitochondria-mediated caspase-3 signaling pathway, and the apoptosis of neuronal cells, thereby exerting the neuroprotective effect.

MiR -18a -5p aggravates homocysteine -induced myocardial injury via autophagy / 中南大学学报(医学版)

OBJECTIVES@#Hyperhomocysteinaemia (Hcy) is an independent risk factor for cardiovascular and cerebrovascular diseases. MicroRNA (miR)-18a-5p is closely related to cardiovascular diseases. This study aims to investigate the effects of miR-18a-5p on homocysteine (Hcy)-induced myocardial cells injury.@*METHODS@#H9c2 cells were transfected with miR-18a-5p mimic/miR-18a-5p mimic negative control (NC) or combined with Hcy for intervention, and untreated cells were set as a control group. The transfection efficiency was verified by real-time RT-PCR, and cell counting kit-8 (CCK-8) assay was used to determine cell viability. Flow cytometry was used to detect apoptosis and reactive oxygen species (ROS) levels. Western blotting was performed to measure the protein levels of microtubule-associated protein 1 light chain 3 (LC3)-I, LC3-II, Beclin1, p62, Bax, Bcl-2, and Notch2. Dual luciferase reporter assay was used to detect the interaction of miR-18a-5p with Notch2.@*RESULTS@#Compared with the control, treatment with Hcy or transfection with miR-18a-5p mimic alone, or combined treatment with Hcy and miR-18a-5p mimic/miR-18a-5p mimic NC significantly reduced the H9c2 cell viability, promoted apoptosis and ROS production, up-regulated the expressions of Bax and Beclin, down-regulated the expressions of Bcl-2, p62, and Notch2, and increased the ratio of LC3-II/LC3-I (all P<0.05). Compared with the combined intervention of miR-18a-5p mimic NC and Hcy group, the above indexes were more significantly changed in the combined intervention of miR-18a-5p mimic and Hcy group, and the difference between the 2 groups was statistically significant (all P<0.05). There is a targeted binding between Notch2 and miR-18a-5p.@*CONCLUSIONS@#MiR-18a-5p could induce autophagy and apoptosis via increasing ROS production in cardiomyocytes, and aggravate Hcy-induced myocardial injury. Notch2 is a target of miR-18a-5p.

LncRNA DRAIC regulates the proliferation, apoptosis, migration and invasion of lung adenocarcinoma cells by targeting let-7i-5p / 中华肿瘤杂志

Objective: To investigate the effects of lncRNA DRAIC on proliferation, apoptosis, migration and invasion of lung adenocarcinoma cells and its mechanism. Methods: Reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR) was used to detect the expression of DRAIC in lung cancer tissues and corresponding adjacent normal tissues of 40 patients with lung adenocarcinoma who underwent surgery in Tangshan People's Hospital from 2019 to 2020. Lung adenocarcinoma cells A549 and H1299 were cultured in vitro and divided into si-NC group, si-DRAIC group, miR-NC group, let-7i-5p mimics group, si-DRAIC+ inhibitor-NC group, and si-DRAIC+ let-7i-5p inhibitor group. CCK-8 method and clone formation experiment were used to detect cell proliferation. Flow cytometry was used to detect cell apoptosis. Transwell array was used to detect the cell migration and invasion. Western blot was used to detect the protein expressions of Caspase-3, Caspase-9, Bcl-2 and Bax. The double luciferase reporter gene experiment was used to verify the regulatory relationship between DRAIC and let-7i-5p. Independent sample t test was used for comparison between two groups, one-way ANOVA was used for comparison between multiple groups, and Pearson correlation analysis was used for correlation analysis. Results: Compared with adjacent tissues, the expression level of DRAIC in lung adenocarcinoma tissues increased (P<0.05), but the expression level of let-7i-5p decreased (P<0.05). The expression levels of DRAIC and let-7i-5p in lung adenocarcinoma tissues were negatively correlated (r=-0.737, P<0.05). The absorbance value of A549 and H1299 cells in the si-DRAIC group at 48, 72 and 96 hours were lower than those in the si-NC group (P<0.05), the number of clones formed [(91.00±6.08 vs. 136.67±6.51); (50.67±1.53 vs. 76.67±4.51)], the number of migration [(606.67±31.34 vs. 960.00±33.06); (483.33±45.96 vs. 741.67±29.67)], the number of invasion [(185.00±8.19 vs. 447.33±22.05); (365.00±33.87 vs. 688.00±32.97)] were lower than those in the si-NC group (P<0.05). However, the apoptosis rates of cells [(13.43±2.79)% vs. (4.53±0.42)%; (23.77±1.04)% vs. (6.60±1.42)%] were higher than those in the si-NC group (P<0.05). The protein expressions of Caspase-3, Caspase-9 and Bax in si-DRAIC group were higher than those in si-NC group, and the protein expression of Bcl-2 was lower than that in si-NC group (P<0.05). DRAIC is located in the cytoplasm. DRAIC targeted and negatively regulated the expression of let-7i-5p. The absorbance values of A549 and H1299 cells in the let-7i-5p mimics group at 48, 72 and 96 hours were lower than those in the miR-NC group (P<0.05), the number of clones formed [(131.33±14.47 vs. 171.33±6.11); (59.33±4.93 vs. 80.33±7.09)], the number of migration [(137.67±3.06 vs. 579.33±82.03); (425.00±11.14 vs. 669.33±21.13)], the number of invasion [(54.00±4.36 vs. 112.67±11.59); (80.00±4.58 vs. 333.33±16.80)] were lower than those in the miR-NC group (P<0.05). However, the apoptosis rates of cells [(14.57±1.10)% vs. (6.97±1.11)%; (23.97±0.42)% vs. (7.07±1.21)%] were higher than those in the miR-NC group (P<0.05). The protein expressions of Caspase-3, Caspase-9 and Bax in let-7i-5p mimics group were higher than those in miR-NC group, and the protein expression of Bcl-2 was lower than that in miR-NC group (P<0.05). The absorbance values of A549 and H1299 cells in the si-DRAIC+ let-7i-5p inhibitor group at 48, 72 and 96 hours were higher than those in the si-DRAIC+ inhibitor-NC group (P<0.05), the number of clones formed [(82.00±5.29 vs. 59.00±5.57); (77.67±4.93 vs. 41.33±7.57)], the number of migration [(774.33±35.81 vs. 455.67±19.04); (569.67±18.72 vs. 433.67±16.77)], the number of invasion [(670.33±17.21 vs. 451.00±17.52); (263.67±3.06 vs. 182.33±11.93)] were higher than those in the si-DRAIC+ inhibitor-NC group (P<0.05). However, the apoptosis rates of cells [(7.73±0.45)% vs. (19.13±1.50)%; (8.00±0.53)% vs. (28.40±0.53)%] were lower than those in the si-NC group (P<0.05). The protein expressions of Caspase-3, Caspase-9 and Bax in si-DRAIC+ let-7i-5p inhibitor group were higher than those in si-DRAIC+ inhibitor-NC group, and the protein expression of Bcl-2 was lower than that in si-DRAIC+ inhibitor-NC group (P<0.05). Conclusion: DRAIC is highly expressed in lung adenocarcinoma, and DRAIC promotes the proliferation, migration and invasion of lung adenocarcinoma cells and inhibits apoptosis by targeting let-7i-5p.

Intermittent heat exposure induces thoracic aorta injury in spontaneously hypertensive rats by activating the AMPK/mTOR/ULK1 pathway / 南方医科大学学报

OBJECTIVE@#To investigate the effects of different manners of heat exposure on thoracic aorta injury in spontaneously hypertensive rats (SHRs) and explore the underlying mechanism.@*METHODS@#Normal 6 to 7-week-old male SHRs were randomized into control group (cage at room temperature), intermittent heat exposure group (SHR-8 group, exposed to 32 ℃ for 8 h daily for 7 days) and SHR-24 group (with continuous exposure to 32 ℃ for 7 days). After the treatments, the pathologies of the thoracic aorta of the rats were observed with HE staining, and the expressions of Beclin1, LC3B and p62 were detected with Western blotting and immunofluorescence assay; TUNEL staining was used to observe cell apoptosis in the thoracic aorta, and the expressions of caspase-3, Bax, and Bcl-2 were detected using Western blotting. The effects of intraperitoneal injections of 3-MA (an autophagy agonist), rapamycin (an autophagy inhibitor) or compound C 30 min before intermittent heat exposure on the expressions of proteins associated with autophagy, apoptosis and the AMPK/mTOR/ULK1 pathway in the aorta were examined with immunohistochemistry.@*RESULTS@#In SHR-8 group, the rats showed incomplete aortic intima with disordered cell distribution and significantly increased expressions of Beclin1, LC3II/LC3I and Bax, lowered expressions of p62 and Bcl-2, and increased apoptotic cells in the thoracic aorta (P < 0.05). Pretreatment with 3-MA obviously inhibited the expressions of autophagy- and apoptosis-related proteins, whereas rapamycin promoted their expressions. Compared with the control group, the rats in SHR-8 group had significantly down-regulated p-mTOR and up-regulated p-AMPK and p-ULK1 expression of in the aorta; Treatment with compound C obviously lowered the expressions of p-AMPK and p-ULK1 and those of LC3B and Beclin1 as well.@*CONCLUSION@#In SHRs, intermittent heat exposure causes significant pathologies and promotes autophagy and apoptosis in the thoracic aorta possibly by activating the AMPK/mTOR/ULK1 pathway.

AAZ2 induces mitochondrial-dependent apoptosis by targeting PDK1 in gastric cancer / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Drastic surges in intracellular reactive oxygen species (ROS) induce cell apoptosis, while most chemotherapy drugs lead to the accumulation of ROS. Here, we constructed an organic compound, arsenical N-‍(4-(1,3,2-dithiarsinan-2-yl)phenyl)acrylamide (AAZ2), which could prompt the ROS to trigger mitochondrial-dependent apoptosis in gastric cancer (GC). Mechanistically, by targeting pyruvate dehydrogenase kinase 1 (PDK1), AAZ2 caused metabolism alteration and the imbalance of redox homeostasis, followed by the inhibition of phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway and leading to the activation of B-cell lymphoma 2 (Bcl2)/Bcl2-associated X (Bax)/caspase-9 (Cas9)/Cas3 cascades. Importantly, our in vivo data demonstrated that AAZ2 could inhibit the growth of GC xenograft. Overall, our data suggested that AAZ2 could contribute to metabolic abnormalities, leading to mitochondrial-dependent apoptosis by targeting PDK1 in GC.

Clinicopathological features and prognosis of high-grade B-cell lymphoma with MYC and bcl-2 and/or bcl-6 rearrangements / 中华病理学杂志

Objective: To investigate the clinicopathological characteristics and prognosis of high-grade B-cell lymphoma (HGBL) involving combined rearrangements of MYC, bcl-2 and bcl-6. Methods: A total of 1 138 cases of large B cell lymphoma (LBL) that were treated at the Ruijin Hospital Affiliated to Shanghai Jiaotong University School of Medicine from January 2017 to September 2020 were analyzed using fluorescence in situ hybridization (FISH) with probes against MYC, bcl-2 and bcl-6. The clinical and pathological data of the 45 patients with HGBL that had rearrangements of MYC and bcl-2 and/or bcl-6 were collected and retrospectively analyzed. Results: Among the 1 138 LBL, 45 (4.0%) cases had combined rearrangements of MYC, bcl-2 and/or bcl-6 that included 6 HGBL cases with MYC, bcl-2 and bcl-6 rearrangements, 14 HGBL cases with MYC and bcl-2 rearrangements, and 25 HGBL cases with MYC and bcl-6 rearrangements. Of these 45 patients, 29 patients were male, and 16 patients were female, aged 29 to 83 years. HGBL with MYC, bcl-2 and bcl-6 rearrangements and HGBL with MYC and bcl-2 rearrangement were reclassified as the germinal center B-cell (GCB) subtype using the Hans algorithm. HGBL with MYC and bcl-6 rearrangement were reclassified as the GCB subtype (68.0%) and the non-GCB subtype (32.0%). The vast majority of HGBL cases had a high Ki-67 proliferation index. Most HGBL patients had advanced stage disease with a high IPI score and an increased LDH level. Also, some patients had clinical features including elevated plasma β2-microglobulin levels, B symptoms, and bone marrow involvement. The IPI scores and LDH levels were significantly different between the HGBL cases with MYC, bcl-2 and bcl-6 rearrangements and the HGBL cases with MYC and bcl-6 rearrangements (P<0.05). Compared with the HGBL cases with MYC, bcl-2 and bcl-6 rearrangements, the HGBL cases with MYC and bcl-2 or bcl-6 rearrangements had a lower incidence of bone marrow involvement (P<0.05). There were no significant differences in the prognosis among HGBL cases with MYC, bcl-2 and bcl-6 rearrangements, the cases with MYC and bcl-2 rearrangements, and the cases with MYC and bcl-6 rearrangements (P>0.05). Conclusions: HGBL with MYC, bcl-2 and/or bcl-6 rearrangements are rare types of B-cell lymphoma with high degree of malignancy and have a short overall survival. To reduce misdiagnosis and improve diagnostic accuracy, it is necessary to assess the patients' clinical features and conduct histopathological, immunohistochemical and FISH analyses.

Suppression of HMGB1 inhibits neuronal autophagy and apoptosis to improve neurological deficits in rats following intracerebral hemorrhage / 南方医科大学学报

OBJECTIVE@#To investigate the effect of suppressing high-mobility group box 1 (HMGB1) on neuronal autophagy and apoptosis in rats after intracerebral hemorrhage (ICH) in rats.@*METHODS@#Rat models of ICH induced by intracerebral striatum injection of 0.2 U/mL collagenase Ⅳ were treated with 1 mg/kg anti-HMGB1 mAb or a control anti-IgG mAb injected via the tail immediately and at 6 h after the operation (n=5). The rats in the sham-operated group (with intracranial injection of 2 μL normal saline) and ICH model group (n=5) were treated with PBS in the same manner after the operation. The neurological deficits of the rats were evaluated using modified neurological severity score (mNSS). TUNEL staining was used to detect apoptosis of the striatal neurons, and the expressions of HMGB1, autophagy-related proteins (Beclin-1, LC3-Ⅱ and LC3-Ⅰ) and apoptosis-related proteins (Bcl-2, Bax and cleaved caspase-3) in the brain tissues surrounding the hematoma were detected using Western blotting. The expression of HMGB1 in the striatum was detected by immunohistochemistry, and serum level of HMGB1 was detected with ELISA.@*RESULTS@#The rat models of ICH showed significantly increased mNSS (P < 0.05), which was markedly lowered after treatment with anti- HMGB1 mAb (P < 0.05). ICH caused a significant increase of apoptosis of the striatal neurons (P < 0.05), enhanced the expressions of beclin-1, LC3-Ⅱ, Bax and cleaved caspase-3 (P < 0.05), lowered the expressions of LC3-Ⅰ and Bcl-2 (P < 0.05), and increased the content of HMGB1 (P < 0.05). Treatment with anti-HMGB1 mAb obviously lowered the apoptosis rate of the striatal neurons (P < 0.05), decreased the expressions of Beclin-1, LC3-Ⅱ, Bax and cleaved caspase-3 (P < 0.05), increased the expressions of LC3-Ⅰ and Bcl-2 (P < 0.05), and reduced the content of HMGB1 in ICH rats (P < 0.05).@*CONCLUSION@#Down- regulation of HMGB1 by anti-HMGB1 improves neurological functions of rats after ICH possibly by inhibiting autophagy and apoptosis of the neurons.