Immune disorders play a vital role in the tumors,infections and allergic diseases.As a co-inhibitory molecule that has attracted much attention in recent years,programmed cell death protein 1(PD-1)can participate in immunomodulation in various immune diseases by inhibiting the proliferation and activation of type 2 innate lymphoid cells(ILC2s).ILC2s is a newly discovered subgroup of immune cell families,which is involved in tumor immunity,anti-parasitic infection and tissue damage repair by secreting type 2 cytokines.PD-1 is a major immunosuppressive molecule that can regulate the different immunomodulatory effects of ILC2s in different immune diseases.This article reviews the role and mechanism of PD-1 in regulating ILC2s in different diseases.

Immune cells are formed by the proliferation and differentiation of hematopoietic stem cells in the bone marrow,in-cluding lymphocytes,macrophages,etc,which participate in the immune defense,immune stability and immune monitoring of the body,and are an indispensable basic component of the immune system.Basic helix-loop-helix family member E40(BHLHE40)has been shown to play an important role in adipogenesis,tumorigenesis,circadian rhythm and hypoxia response.Recently,it was found that BHLHE40 plays a key role in the cell cycle,proliferation and cytokine production of immune cells.This article will review the discovery,naming,structure,and mechanism of BHLHE40 in various periods,as well as it's research progress in immune cells,in order to provide new target for the treatment of immune-related diseases at the genetic level.

Immune disorders play a vital role in the tumors,infections and allergic diseases.As a co-inhibitory molecule that has attracted much attention in recent years,programmed cell death protein 1(PD-1)can participate in immunomodulation in various immune diseases by inhibiting the proliferation and activation of type 2 innate lymphoid cells(ILC2s).ILC2s is a newly discovered subgroup of immune cell families,which is involved in tumor immunity,anti-parasitic infection and tissue damage repair by secreting type 2 cytokines.PD-1 is a major immunosuppressive molecule that can regulate the different immunomodulatory effects of ILC2s in different immune diseases.This article reviews the role and mechanism of PD-1 in regulating ILC2s in different diseases.

Immune cells are formed by the proliferation and differentiation of hematopoietic stem cells in the bone marrow,in-cluding lymphocytes,macrophages,etc,which participate in the immune defense,immune stability and immune monitoring of the body,and are an indispensable basic component of the immune system.Basic helix-loop-helix family member E40(BHLHE40)has been shown to play an important role in adipogenesis,tumorigenesis,circadian rhythm and hypoxia response.Recently,it was found that BHLHE40 plays a key role in the cell cycle,proliferation and cytokine production of immune cells.This article will review the discovery,naming,structure,and mechanism of BHLHE40 in various periods,as well as it's research progress in immune cells,in order to provide new target for the treatment of immune-related diseases at the genetic level.

Sepsis is a life-threatening organ dysfunction caused by the imbalance of host immune response to infection,which immune regulation mechanism is complex and has not been clarified.Programmed death receptor-1(PD-1)/programmed death recep-tor ligand-1(PD-L1)is a negative co-inhibitory molecule that has attracted much attention in recent years.Studies have shown that PD-1/PD-L1 signaling pathway plays an important immunosuppressive role in sepsis.There are differences in immune regulation mecha-nisms in heart,liver,spleen,lung and kidney,which has not been clarified.Immune regulation of PD-1/PD-L1 in different organs of sepsis is reviewied in this article,which could to provide a new direction for the study of sepsis.

Objective: To investigate whether microRNA(miR)-214 can improve hyperglycemia induced pyroptosis in H9c2 cells through targeting caspase-1. Methods: H9c2 cells of rats those in good growth condition were selected and incubated into the T25 culture bottle after digestion and passage. Cells were cultured in an incubator at 37 ℃ with 5%CO₂, repeat passage was made after cell density reached about 80%, The 5^th to 8^th generations of cells were selected for the subsequent experiments. To observe the effect of overexpression of miR-214 on pyroptosis and caspase-1 expression in H9c2 cells induced by hyperglycemia, the cells were divided into 4 groups: Control group(H9c2 cells cultured normally), Hyperglycemia group (HG group, 50 mmol/L glucose was used to intervene H9c2 cells for 24 hours), miR-214 mimics+hyperglycosis group (mimics+HG group, H9c2 cells were transfected with miR-214 mimics for 24 hours and then treated with 50 mmol/L hyperglycosis for 24 hours), miR-214 mimic-negative control+hyperglycaemic group(MNC+HG group, H9c2 cells were transfected with miR-214 mimic-negative control for 24 hours and then treated with 50 mmol/L hyperglycaemic for 24 hours). In order to further verify the anti-pyroptosis effect of miR-214 was mediated by targeted inhibition on caspase-1, cells overexpressing caspase-1 were used in the rescue experiment. The cells overexpressing caspase-1 were divided into 4 groups: Hyperglycemia group (HG group, 50 mmol/L glucose was used to intervene H9c2 cells for 24 hours), miR-214 mimics+hyperglycosis group (mimics+HG group, H9c2 cells were transfected with miR-214 mimics for 24 hours and then treated with 50 mmol/L hyperglycosis for 24 hours), miR-214 mimics+hyperglycosis+recombinant adenovirus (Ad-caspase-1-EGFP) group with caspase-1 gene and EGFP green fluorescent protein expression (mimics+HG+Ad-caspase-1-EGFP group, H9c2 cells were transfected with caspase-1-green fluorescent protein-carrying adenovirus for 48 hours, followed by transfection of miR-214 mimics for 24 hours, and then treated with 50 mmol/L hyperglycaemia for 24 hours), miR-214 mimics+HG+Ad-EGFP empty virus group (mimics+HG+Ad-EGFP group, H9c2 cells were transfected with empty adenovirus containing green fluorescent protein for 48 hours, followed by transfection with miR-214 mimics for 24 hours, and then treated with 50 mmol/L hyperglycosis for 24 hours). The mRNA expression levels of miRNA-214 and caspase-1 in cells were detected by real-time quantitative PCR. The expression and localization of caspase-1 protein were detected by immunofluorescence assay. Western blot was used to detect protein expression levels of procaspase-1, cleaved caspase-1, NLRP3 and ACS with β-actin as internal reference. The secretion of IL-1β and IL-18 in cell culture medium was detected by ELISA. The correlation between miR-214 and caspase-1 was detected by double luciferase reporter gene. Results: (1) The mRNA expression levels of miR-214 and caspase-1 in each group: the mRNA expressions of miR-214 in HG group and MNC+HG group were significantly lower than that in control group(P<0.05). The mRNA expression of miR-214 in mimics+HG group was significantly higher than that in control group (P<0.05). The mRNA expression levels of caspase-1 in HG group and MNC+HG group were significantly higher than that in control group(P<0.05). The mRNA expression level of caspase-1 in mimics+HG group was lower than that in control group(P<0.05). (2) The expression of caspase-1 in each group: the green fluorescence intensity in the control group was weak, which was strong in the HG group and MNC+HG group. The green fluorescence expression was weaker in mimics+HG group than in HG group. (3) ASC and NLRP3 protein expression levels in each group: ASC and NLRP3 protein expression levels in HG group and MNC+HG group were higher than those in control group(P<0.05). ASC and NLRP3 protein expression levels were significantly lower in mimics+HG group than in mimics+HG group (P<0.05). (4) The secretion of IL-1β and IL-18 in the cell culture medium of each group: the content of IL-1β and IL-18 in the cell culture medium of HG group and MNC+HG group was significantly higher than that of control group (P<0.05). The content of IL-1β and IL-18 in the cell culture medium of mimics+HG group was significantly lower than that of the HG group (P<0.05). (5) Correlation between miR-214 and caspase-1: miR-214 specifically binds to caspase-1 3 ′UTR. Meanwhile, Western blot results showed that cleaved caspase-1 protein expression levels were significantly higher in both HG group and MNC+HG group than in control group (P<0.05). The levels of cleaved caspase-1 were significantly lower in mimics+HG group than in HG group (P<0.05). There was no significant difference in procaspase-1 expression among groups (P>0.05). (6) The expression levels of procaspase-1, cleaved caspase-1, ASC and NLRP3 in each group in rescue experiment: there was no significant difference in the expression of procaspase-1 in each group (P>0.05). Cleaved caspase-1, ASC and NLRP3 protein expressions were significantly lower in mimics+HG group than in HG group (P<0.05). However, cleaved caspase-1, ASC and NLRP3 protein expressions were significantly higher in mimics+HG+ Ad-caspase-1-EGFP group than in mimics+HG group (P<0.05). (7) The expression of IL-1β and IL-18 in rescue experiment: the secretions of IL-1β and IL-18 in the cell culture medium of the mimics+HG group were significantly lower than that of HG group (P<0.05), which were significantly higher in mimics+HG+Ad-caspase-1-EGFP group than in mimics+HG group (P<0.05). Conclusion miR-214 can improve the hyperglycemia induced pyroptosis in H9c2 cells by targeted inhibition of the caspase-1.

Objective To investigate whether microRNA(miR)?214 can improve hyperglycemia induced pyroptosis in H9c2 cells through targeting caspase?1. Methods H9c2 cells of rats those in good growth condition were selected and incubated into the T25 culture bottle after digestion and passage. Cells were cultured in an incubator at 37℃with 5%CO2, repeat passage was made after cell density reached about 80%, The 5th to 8th generations of cells were selected for the subsequent experiments. To observe the effect of overexpression of miR?214 on pyroptosis and caspase?1 expression in H9c2 cells induced by hyperglycemia, the cells were divided into 4 groups: Control group(H9c2 cells cultured normally), Hyperglycemia group (HG group, 50 mmol/L glucose was used to intervene H9c2 cells for 24 hours), miR?214 mimics+hyperglycosis group (mimics+HG group, H9c2 cells were transfected with miR?214 mimics for 24 hours and then treated with 50 mmol/L hyperglycosis for 24 hours), miR?214 mimic?negative control+hyperglycaemic group(MNC+HG group, H9c2 cells were transfected with miR?214 mimic?negative control for 24 hours and then treated with 50 mmol/L hyperglycaemic for 24 hours). In order to further verify the anti?pyroptosis effect of miR?214 was mediated by targeted inhibition on caspase?1, cells overexpressing caspase?1 were used in the rescue experiment. The cells overexpressing caspase?1 were divided into 4 groups: Hyperglycemia group (HG group, 50 mmol/L glucose was used to intervene H9c2 cells for 24 hours), miR?214 mimics+hyperglycosis group (mimics+HG group, H9c2 cells were transfected with miR?214 mimics for 24 hours and then treated with 50 mmol/L hyperglycosis for 24 hours), miR?214 mimics+hyperglycosis+recombinant adenovirus (Ad?caspase?1?EGFP) group with caspase?1 gene and EGFP green fluorescent protein expression (mimics+HG + Ad?caspase?1?EGFP group, H9c2 cells were transfected with caspase?1?green fluorescent protein?carrying adenovirus for 48 hours, followed by transfection of miR?214 mimics for 24 hours, and then treated with 50 mmol/L hyperglycaemia for 24 hours), miR?214 mimics+HG+Ad?EGFP empty virus group (mimics+HG+Ad?EGFP group, H9c2 cells were transfected with empty adenovirus containing green fluorescent protein for 48 hours, followed by transfection with miR?214 mimics for 24 hours, and then treated with 50 mmol/L hyperglycosis for 24 hours). The mRNA expression levels of miRNA?214 and caspase?1 in cells were detected by real?time quantitative PCR. The expression and localization of caspase?1 protein were detected by immunofluorescence assay. Western blot was used to detect protein expression levels of procaspase?1, cleaved caspase?1, NLRP3 and ACS with β?actin as internal reference. The secretion of IL?1β and IL?18 in cell culture medium was detected by ELISA. The correlation between miR?214 and caspase?1 was detected by double luciferase reporter gene. Results (1) The mRNA expression levels of miR?214 and caspase?1 in each group: the mRNA expressions of miR?214 in HG group and MNC+HG group were significantly lower than that in control group(P<0.05). The mRNA expression of miR?214 in mimics+HG group was significantly higher than that in control group (P<0.05). The mRNA expression levels of caspase?1 in HG group and MNC+HG group were significantly higher than that in control group(P<0.05). The mRNA expression level of caspase?1 in mimics+HG group was lower than that in control group(P<0.05). (2) The expression of caspase?1 in each group: the green fluorescence intensity in the control group was weak, which was strong in the HG group and MNC+HG group. The green fluorescence expression was weaker in mimics+HG group than in HG group. (3) ASC and NLRP3 protein expression levels in each group: ASC and NLRP3 protein expression levels in HG group and MNC+HG group were higher than those in control group(P<0.05). ASC and NLRP3 protein expression levels were significantly lower in mimics+HG group than in mimics+HG group (P<0.05). (4) The secretion of IL?1β and IL?18 in the cell culture medium of each group: the content of IL?1β and IL?18 in the cell culture medium of HG group and MNC+HG group was significantly higher than that of control group (P<0.05). The content of IL?1β and IL?18 in the cell culture medium of mimics+HG group was significantly lower than that of the HG group (P<0.05). (5) Correlation between miR?214 and caspase?1: miR?214 specifically binds to caspase?1 3 ＇UTR. Meanwhile, Western blot results showed that cleaved caspase?1 protein expression levels were significantly higher in both HG group and MNC+HG group than in control group (P<0.05). The levels of cleaved caspase?1 were significantly lower in mimics+HG group than in HG group (P<0.05). There was no significant difference in procaspase?1 expression among groups (P>0.05). (6) The expression levels of procaspase?1, cleaved caspase?1, ASC and NLRP3 in each group in rescue experiment: there was no significant difference in the expression of procaspase?1 in each group (P>0.05). Cleaved caspase?1 , ASC and NLRP3 protein expressions were significantly lower in mimics+HG group than in HG group (P<0.05). However, cleaved caspase?1, ASC and NLRP3 protein expressions were significantly higher in mimics+HG+Ad?caspase?1?EGFP group than in mimics+HG group (P<0.05). (7) The expression of IL?1β and IL?18 in rescue experiment: the secretions of IL?1β and IL?18 in the cell culture medium of the mimics+HG group were significantly lower than that of HG group (P<0.05), which were significantly higher in mimics+HG+Ad?caspase?1?EGFP group than in mimics+HG group (P<0.05). Conclusion miR?214 can improve the hyperglycemia induced pyroptosis in H9c2 cells by targeted inhibition of the caspase?1.

Objective: To explored the effect and related mechanism of miRNA-21 on hydrogen peroxide-induced C-kit⁺ cardiac stem cells apoptosis. Methods: C-kit⁺ cardiac stem cells were isolated from SD rats by the methods of enzyme digestion and magnetic bead. Cells were divided into the following experimental groups: (1) negative control mimics (NCM)group: cells were transfected with negative control miRNA-21 mimics for 48 hours; (2)mimics group: cells were transfected with miRNA-21 mimics for 48 hours; (3) NCM+ H₂O₂ group: negative control miRNA-21 mimics were transfected into cells for 48 hours and then treated with 100 μmol H₂O₂ for 2 hours; (4)mimics+ H₂O₂ group: miRNA-21 mimics were transfected into cells for 48 hours and then treated with 100 μmol H₂O₂ for 2 hours. mRNA of miRNA-21 was detected by RT-PCR. The apoptosis rate of C-kit⁺ cardiac stem cells was determined using the annexin V-FITC/PI staining assay. Western blot was employed to measure the expression of apoptosis related proteins(Caspase-3, Bax, and Bcl-2). Results: (1) Compared with the NCM group, the mRNA expression level of miRNA-21 was significantly up-regulated in mimics group, while obviously down-regulated in NCM+ H₂O₂ group(all P<0.05). Compared with the mimics group, the mRNA expression levels of miRNA-21 in mimics+ H₂O₂ group was significantly downregulated (P<0.05), but remarkably upregulated compared with the NCM+ H₂O₂ group(P<0.05). (2) Flow cytometry results indicated that the early apoptosis rates were similar between the NCM group and mimics group ((4.57±3.45)% vs. (5.13±3.21)%, P>0.05). Compared with the NCM group, the early apoptosis rates were remarkably increased ((79.07±5.75)% vs.(4.57±3.45)%, P<0.05) in NCM+ H₂O₂ group. Compared with the mimics group, the early apoptosis was significantly up-regulated in the mimics+ H₂O₂ group ((30.27±1.36)% vs.(5.13±3.21)%, P<0.05), which were further down-regulated in mimics+ H₂O₂ group compared with the NCM+ H₂O₂ group ((30.27±1.36)% vs.(79.07±5.75)%, P<0.05). (3) Western blot results showed similar protein expression of Caspase-3, Bax and Bcl-2 between NCM group and mimics group(all P>0.05). Compared with the NCM group, the Caspase-3 and Bax protein expression was significantly increased in NCM+ H₂O₂ group (all P<0.05), but the protein expression level of Bcl-2 was similar between the 2 groups(P>0.05). The Caspase-3 and Bax protein expression was markedly decreased, while Bcl-2 apparently increased in the mimics+ H₂O₂ group compared with the NCM+ H₂O₂ group(all P<0.05). Conclusion Overexpression of miRNA-21 protects the C-kit⁺ cardiac stem cells from apoptosis caused by oxidative stress through downregulating proapoptotic and upregulating the antiapoptotic proteins.

AIM:To investigate the effect of calcitonin gene-related peptide (CGRP)transfection into c-kitpos cardiac stem cells (c-kit+CSCs) on the cell viability.METHODS: Under the sterile condition, the auricles of SD rats were taken out , and then c-kit +CSCs were collected through enzyme digestion and immunomagnetic bead separation (MACS).The cells were identified by flow cytometry .c-kit +CSCs were transfected with enhanced green fluorescent pro-tein CGRP lentiviral vector ( Lv-EGFP-CGRP) or enhanced green fluorescent protein lentiviral vector ( Lv-EGFP).The cells were randomly divided into Lv-EGFP-CGRP-CSCs group , Lv-EGFP-CSCs group and CSCs group .The transfection was observed under the fluorescence microscope .The transfection efficiency was detected by flow cytometry .The CGRP protein secretion in the cell culture supernatants was detected by ELISA .The viability of c-kit+CSCs transfected with Lv-EGFP-CGRP or Lv-EGFP was measured by CCK-8 assay.RESULTS:c-kit+CSCs were isolated and cultured successfully .The expression positive rate of c-kit was 91.0%and the expression positive rates of CD 45 and CD34 were 4.5% and 4.0%, respectively.After transfected with lentivirus for 48 h, the stable fluorescence in c-kit +CSCs was observed under fluores-cence microscope .The transfection efficiency were 80%when MOI was 20.The level of CGRP was significantly increased in Lv-ECFP-CGRP-CSCs group compared with Lv-EGFP-CSCs group and CSCs group (P<0.05).Meanwhile, transfection with lentiviral vector in each group did not affect the viability of c-kit+CSCs.CONCLUSION:Transfection of Lv-EGFP-CGRP into c-kit+CSCs was successful .The secretion of CGRP was found in the transfected c-kit+CSCs and the viability was not changed after transfection .CGRP-modified c-kit+CSCs may play a role in treating myocardial infarction .

Objective: Pocket infection in patients after total removal of implanted pacemaker has many problems for their electronic system; our research intends to explore the feasibility of conservatively treating such infection and retain the electronic system. Methods: A total of 4 patients with pacemaker pocket infection in our hospital from 2015-01 to 2016-02 were studied. Thorough debridement and disinfection were conducted in infected pockets and devices, meanwhile vacuum sealing drainage was applied. Electrode wire was kept and intravenous antibiotics were given for (7-10) days after the operation in all patients. Results: The average time of infection occurred at 14.75 months after operation with the type of isolated pacemaker pocket infection. Pocket vacuum suction drainage was performed, with the mean of 7.25 (5-10) months follow-up observation, infection was disappeared and the patients had good wound healing. Conclusion: With thorough debridement of infected pocket, rational treatment of residual electronic system and vacuum sealing drainage, the infection might be effectively controlled for complete recovery without lead extraction in relevant patients.