It is proposed that following peripheral nerve injury abnormal sprouting of Aβ fiber primary afferent neurons in the spinal cord contributes to the allodynia that often occurs with such injury. The present investigation is to determine whether this sprouting is reversal after compression of peripheral nerve was relieved. In a rat model of neuropathic pain made by rat sciatic nerve compression,chorela toxin B subunit conjugated horseradish peroxidase (CB-HRP) was used to trace the termination of Afiber primary afferents and sections were reacted for using tetramethylbenzidine (TMB) as the chromagen. We demonstrated that the compression to the sciatic nerve also results in hyperalgesia and novel transganglionic CB-HRP staining in laminae Ⅱ, and this sprouting can not be reversed by decompression. This structural reorganization in central nervous system and its irreversible character may contribute to the development and maintenance of neuropathic pain.

OBJECTIVE:To study the absorptive characteristics of ginsenoside Rb3 in Caco-2 cell monolayer model. METHO-DS:The ginsenoside Rb3 cell samples underwent high speed centrifugation, then the supernatant was collected and determined by LC-ESI-MS/MS method in which the mobile phase consisted of acetonitrile-1 mmol?L-1 ammonium formate water solution (34 ∶ 66) with ginsenoside Rg2 as internal standard, and the tandem mass spectrometry was operated in negative electrospray ionization in a multiple reaction monitoring (MRM) mode, with detection ions m/z1077.7→m/z 783.4 for ginsenoside Rb3 and m/z 783.6→m/z 475.1 for ginsenoside Rg2.The concentration of ginsenoside Rb3 across the Caco-2 cell monolayer model was determined and the apparent permeability coefficient(Papp) of ginsenoside Rb3 was calculated. RESULTS: The calibration curve for ginsenoside Rb3 was linear in the range of 50～2 000 ng?mL-1,with intra-day precision and inter-day precision at less than 15%. P(AP-BL) from apical side (AP) to basolateral side (BL) was 3.22?10-6 cm?s-1, whereas P(BL-AP) from BL to AP was 6.0?10-6 cm?s-1,and the ratio of P(BL-AP)/P(AP-BL) was 1.86.CONCLUSION:The LC-ESI/MS/MS method is simple and sensitive, and it is applicable for the study of the absorptive characteristics of ginsenoside Rb3 in Caco-2 cell monolayer model.

Objective To evaluate the development of fetal cerebellar vermis using 3D transabdominal ultrasound,and provide evidence for prenatal screening fetal cerebellar vermis anomalies.Methods Totally 387 normal fetuses at 20～36 gestation weeks were examined by three-dimensional extended imaging(3DXI) to observe the fetal cerebellar vermis.The width,anteroposterior diameter,craniocaudal diameter of the cerebellar vermis,the angle between brain stem and vermis,a ratio between the area of anterior vermis and posterior vermis were measured,and the relationship between the gestational age and parameters mentioned above were analyzed.Results The multi slice view of 3DXI and 3D reconstructed sagittal section well evaluated the integrity of vermis morphous and vermis size,identified characteristic signs of vermis:the fourth ventricle apex and vermis crack.In normal fetus,brain stem and vermis were almost parallel,the angle between them was 3.97°±1.65°.There was no significant correlation between the angle and the gestational age.The area of the anterior vermis was smaller than that of posterior vermis,with a ratio of 0.76±0.06,which was also not related to gestational age.The width,anteroposte rior diameter and craniocaudal diameter of the cerebellar vermis were positively correlated with gestational age.Conclusions The multi slice view of 3DXI and 3D reconstructed sagittal section should help evaluate the development of the cerebellar vermis,accurately show the cerebellar vermis and its surrounding strutures,and provide a new way to evaluate the fetal cerebellar vermis.

Objective:To investigate the role of growth arrest specific protein 6 (Gas6) in regulating macrophage polarization in wound healing.Methods:Clean male B6 mice were randomly(random number) divided into the normal group, skin defect group, skin defect group + normal saline group (PBS group), skin defect + Gas6 (1 μg) group, skin defect + Gas6 (5 μg) group, and skin defect + Gas6 (10 μg) group. Ten mice in each group were used to observe the healing of skin wounds. Macrophages were isolated from the wound tissues of the remaining 6 mice on the fifth day after modeling. The levels of IL-6 and IL-10 were detected by enzyme-linked immunosorbent assay (ELISA), the mRNA expression levels of arginase-1 (Arg-1) and inducible nitric oxide synthase (iNOS) were detected by RT-PCR, and flow cytometry was used to detect the expression of M1 marker CD197, M2 marker CD163 and F4/80. HE staining was used to detect the pathological changes of skin wounds. Masson staining was used to analyze the granulation tissue and collagen deposition.Results:Scab began to form on the surface of the wound on the third day after the skin defect model was established. The wound area of the Gas6 treatment group was smaller than that of the PBS group, and the wound healing was better than that of the PBS group. Compared with the normal group, the proportion of CD197 in macrophages of the skin defect group was significantly increased ( P=0.00 49), the proportion of CD163 and F4/80 double positive was significantly decreased ( P=0.00 86), the level of IL-6 was significantly increased ( P=0.00 13), the level of IL-10 was significantly increased ( P=0.00 14), the level of iNOS mRNA was significantly increased ( P=0.00 8), and Arg-1 was significantly increased in the skin defect group The mRNA level was significantly decreased ( P=0.01 21), and the inflammatory infiltration was aggravated. Compared with the PBS group, the proportion of CD197 in the Gas6 treatment group was significantly decreased ( P=0.00 0), the double positive rates of CD163 and F4/80 were significantly increased ( P = 0.00 0), the level of IL-6 was significantly decreased (P = 0.00 0), the level of IL-10 was significantly increased ( P=0.00 03), the level of iNOS mRNA was significantly decreased ( P=0.00 18), the level of Arg-1 mRNA was significantly increased ( P=0.00 1), and the number of inflammatory cells and the number of collagen fibers were increased. Conclusions:Gas6 can promote the transformation of macrophages from M1 to M2 in mice with skin defect, which is beneficial to the wound healing of skin defect.

Objective:To explore the prognostic factors of patients with Vibrio vulnificus sepsis. Methods:The clinical data of 67 patients with Vibrio vulnificus sepsis from January 2008 to December 2019 in the First Affiliated Hospital of Wenzhou Medical University were retrospectively analyzed. Univariate analysis was used to compare the differences in general information, clinical manifestations, admission laboratory indicators, antibiotics and surgery between the death group and the cured group. Then the factors with significant difference in univariate analysis were included in multivariate analysis, and the factors of prognosis were obtained. Results:Univariate analysis showed that there were significant difference in liver disease, admission with hypotension shock, multiple limb injuries; admission leukocytes, platelets, pH value, albumin, lactic acid, aspartate aminotransferase, creatinine, procalcitonin, creatine kinase, activated partial thromboplastin time, prothrombin time between the death group and the cured group (all P <0.05). Multivariate analysis showed that admission lactate ( OR=0.628, 95% CI: 0.461-0.855, P=0.003), albumin ( OR=1.330, 95% CI:1.062-1.667, P=0.013), creatine kinase ( OR=0.999, 95% CI: 0.998-1.000, P=0.016) and admission surgery time ( OR=0.118, 95% CI: 0.015-0.938, P=0.043) were risk factors of the prognosis. Patients with high lactate, creatine kinase and low albumin at admission indicate poor prognosis; patients with admission surgery time≤ 12 h have better prognosis. Conclusion:For the treatment of patients with Vibrio vulnificus sepsis, medical staff should dynamically evaluate these prognostic factors in the early stage, and early surgical treatment should be adopted to improve the prognosis of patients.

Objective To investigate the effect of resveratrol (Res) on paraquat (PQ)-induced acute lung injury (ALI) and mortality in mice and the mechanism of nuclear factor-κB (NF-κB) inflammatory pathway. Methods Sixty-eight healthy male ICR mice with grade SPF were enrolled, among them 20 mice were used for mortality observation (n = 10), and other 48 were used for determination of related parameters (n = 6). The mice were randomly divided into four group s: normal saline (NS) control group, Res control group, PQ group and PQ + Res group. The mice in the latter two groups were subdivided into 6, 24, 72 hours subgroups. The PQ poisoning model of mice was reproduced by one injection of 30 mg/kg PQ intraperitoneally. The mice in PQ + Res group were given 60 mg/kg Res intraperitoneally on the contralateral side after PQ injection. The mice were sacrificed at 6, 24, 72 hours after PQ poisoning, and lung tissue was harvested. The serum levels of tumor necrosis factor-α (TNF-α), interleukins (IL-6 and IL-1β) were determined by enzyme linked immunosorbent assay (ELISA). The pathological changes in lung tissue were observed with electron microscopy. Apoptosis cells in the lung were identified by terminal dexynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) for the estimation of apoptosis rate. The protein expression of NF-κB p65 was determined by Western Blot. Results Compared with PQ group, the death number of mice at 48, 72, 96 hours in PQ + Res group was slightly decreased (0 vs. 2, 2 vs. 5, 4 vs. 6) but without statistically significant difference (all P > 0.05). Under electron microscope, the lung injury in PQ group was severer than that in NS control group, and Res was found to be able to alleviate the lung injury. Compared with NS control group [(2.45±0.61)%], the apoptosis rate at 6 hours in PQ group was significantly increased [(8.42±1.48)%], and peaked at 72 hours [(21.23±3.47)%]. Res could decrease the apoptosis rate after PQ poisoning [6 hours: (5.56±1.31)% vs. (8.42±1.48)%, 24 hours: (11.14±2.07)% vs. (16.88±2.96)%, 72 hours: (13.28±2.32)% vs. (21.23±3.47)%, all P < 0.05]. The serum levels of TNF-α, IL-6, and IL-1β, and NF-κB p65 in lung tissue were all markedly increased after PQ poisoning, and they were significantly decreased after Res intervention as compared with those of PQ group [TNF-α (ng/L): 2.62±0.29 vs. 4.06±0.74 at 6 hours, 3.98±0.41 vs. 6.79±0.80 at 24 hours, 5.06±0.75 vs. 11.00±0.75 at 72 hours; IL-6 (ng/L): 14.19±1.54 vs. 16.55±1.24 at 6 hours, 13.21±1.37 vs. 19.73±0.85 at 24 hours, 13.72±0.56 vs. 22.45±0.72 at 72 hours; IL-1β (ng/L): 8.54±1.64 vs. 12.59±0.66 at 6 hours, 10.15±0.29 vs. 16.24±1.03 at 24 hours, 16.14±0.70 vs. 19.55±0.56 at 72 hours; 6-hour NF-κB p65: (1.34±0.07) folds vs. (1.86±0.11) folds when the expression in NS control group was represented as 1, all P < 0.05]. Conclusions Res cannot lower the mortality in mice with PQ poisoning, but it seems to be able to attenuate PQ-induced ALI and cell apoptosis. The mechanism responsible for the latter maybe the inhibitive effect of Res on NF-κB p65 translocation and cytokines production.

Objective To evaluate the relationship between changes in B-type natriuretic peptide(BNP) and cardiac troponin I(cTnI)levels and prognosis of critically ill patients with sepsis. Methods This study retrospectively reviewed the clinical data of 75 patients with severe sepsis and septic shock admitted into Emergency Intensive Care Unit(EICU)of the First Affiliated Hospital of Wenzhou Medical University in Zhejiang Province. According to the severity of the cases,they were divided into two groups:severe sepsis group(34 cases)and septic shock group(41 cases),and based on the difference in prognosis,they were divide into survivor group(32 cases) and non-survivor group(43 cases). Electrocardiogram(ECG)was performed within 24 hours after admission in all the patients. Acute physiology and chronic health evaluation Ⅱ(APACHEⅡ)score and biochemical markers showing organ dysfunctions as BNP, cTnI, creatine kinase (CK), creatine kinase MB mass(CK-MB), and lactate were compared between severe sepsis and septic shock groups and between survivor and non-survivor groups. Results The septic shock group had significantly higher baseline BNP,cTnI,lactate and APACHE Ⅱscore and mortality rate than those in severe sepsis group〔BNP(μg/L):1.90(1.08,2.79)vs. 0.41(0.31,0.75),cTnI (μg/L):1.15(0.92,1.28)vs. 0.58(0.40,0.79),lactate(mmol/L):6.63±3.72 vs. 3.28±1.66,APACHEⅡscore:26.00(24.00,28.00)vs. 21.50(20.00,29.25),mortality rate:70.73%vs. 41.18%,P<0.05 or P<0.01〕. Compared with survivor group,the ages of non-survivor group were older with more males and higher BNP,cTnI,lactate and APACHEⅡscore〔males(cases):30 vs. 13,age(years old):66.49±14.97 vs. 58.19±17.05,BNP:1.60(0.62, 2.51)vs. 0.57(0.37,1.79),lactate:4.10(3.00,9.00)vs. 3.10(2.13,4.18),cTnI:1.02±0.49 vs. 0.62±0.37, APACHE Ⅱ score:28.00(25.00,30.00)vs. 21.00(20.00,25.75),P<0.05 or P<0.01〕. However,there were no statistically significant differences in the levels of CK and CK-MB between the above compared groups(both P>0.05). The patients' ECGs had no obvious changes. Conclusions High plasma BNP and cTnI levels in patients with sepsis may suggest myocardial damage and relatively bad prognosis. The examination of BNP and cTnI levels may help clinicians to early detect the high-risk patients with septic cardiac dysfunction and assess their prognoses.

AIM:To investigate the effect of capsaicin on lipopolysaccharide ( LPS)-induced activation of cul-tured endothelial cells of mouse aorta in vitro.METHODS:The endothelial cells were isolated from mouse aorta and cul-tured in vitro, and the specific cell markers of the cells were identified by immunofluorescence staining.The cells were stimulated with LPS (100μg/L) combined with or without capsaicin, and the cells and supernatant were collected at 12 h, 24 h and 48 h.The levels of soluble intercellular adhesion molecule 1 ( sICAM-1) , soluble vascular cell adhesion molecule 1 (sVCAM-1) and soluble P-selectin (sP-selectin) in the supernatant were measured by ELISA.The levels of nuclear NF-κB p65 and cytopasmic p-IκBαand IκBαwere detected by Western blotting.RESULTS: Compared with control group, the levels of sP-selectin, sICAM-1 and sVCAM-1 in LPS group were significantly increased (P<0.05), and LPS promoted the expression of sICAM-1 and sVCAM-1 in a time-dependent manner.Compared with LPS group at the same time point, capsaicin inhibited the expression of sP-selectin, sICAM-1 and sVCAM-1 in a dose-dependent manner.Compared with con-trol group, the protein levels of NF-κB p65 and p-IκBαin LPS group at 24 h were significantly increased (P<0.05), while the protein level of IκBαin LPS group at 24 h were significantly decreased (P<0.05).Compared with LPS group, capsaicin decreased the protein levels of NF-κB p65 and p-IκBαand increased the protein level of IκBαin a dose-depend-ent manner.CONCLUSION:Capsaicin has a protective effect on LPS-induced vascular endothelial cell activation, which potentially contributes to the suppression of IκBαdegradation and NF-κB p65 nuclear translocation.

Objective To observe the release of glutamate (Glu)and γ-amino butyric acid (GABA) from PC 12 cells induced by aconitine,and to study the intervention of Shuanghuanglian on the injury of these cells. Methods The cell proliferation test agent in cell counting kit(CCK-8)was applied to assay the aconitine toxicity to PC12 cells and to establish the PC12 cell injury model induced by aconitine. The PC12 cells during logarithmic growing phase were randomly divided into the following groups:blank control group(complete medium containing 0.1% dimethyl sulfoxide was added), Shuanghuanglian control group (complete medium containing 50 μg/mL Shuanghuanglian),baicalin control group(complete medium containing 20 μmol/L baicalin),aconitine toxic group(complete medium containing 100 μmol/L aconitine),Shuanghuanglian intervention group(complete medium containing 100μmol/L aconitine and 50μg/mL Shuanghuanglian)and baicalin intervention group(complete medium containing 100 μmol/L aconitine and 20 μmol/L baicalin). The cells in all groups were incubated for 24 hours respectively. The changes of PC12 cell absorbance(A)values were detected by CCK-8 assay before and after intervention by Shuanghuanglian and baicalin. The PC12 cell apoptosis was determined by flow cytometry. Glu and GABA contents in cell culture medium were determined by chromatometry and enzyme-linked immunosorbent assay (ELISA). Results Compared with blank control group,after the PC12 cells treated with 100 μmol/L aconitine for 24 hours,their cytoactivity was decreased markedly(A value:1.003±0.042 vs. 1.685±0.030,P<0.05),then afterwards in the experiment,the incubation of 100 μmol/L aconitine with PC12 cells for 24 hours was considered as the intervention concentration. In blank control group,the normal PC12 cells accounted for 95.89%,while in the aconitine toxic group,the rate of injured PC12 cells reached 64.27% and early apoptosis rate reached 45.46%, and in Shuanghuanglian intervention group and baicalin intervention group,the early apoptosis rate was decreased to 33.24% and 28.22% respectively. Compared with blank control group,there were no significant differences in cytoactivities and the contents of Glu and GABA released by PC12 cells in Shuanghuanglian control group and baicalin control group(all P<0.05),while in the aconitine toxic group,the cytoactivity was significantly decreased(A value:1.056±0.039 vs. 1.722±0.083),and the contents of Glu and GABA were significantly increased〔Glu(μmol/L):5.295±0.137 vs. 3.433±0.138;GABA(μmol/L):0.769±0.020 vs. 0.528±0.012,both P<0.05〕. Compared with aconitine toxic group,the cytoactivities of PC12 were significantly elevated(1.202±0.059 and 1.180±0.032),the levels of Glu were significantly reduced(4.055±0.086 and 3.984±0.057),and the contents of GABA were obviously increased(0.809±0.016 and 0.930±0.021)in the cell culture medium of the Shuanghuanglian intervention group and baicalin intervention group(all P<0.05). The increase of cytoactivity in Shuanghuanglian intervention group was more marked than that of baicalin intervention group(P<0.05). There were no statistical significant differences in contents of Glu and GABA between Shuanghuanglian intervention group and baicalin intervention group(both P>0.05). Conclusions The changes of Glu and GABA may be one of the mechanisms of neural toxic effect of aconitine. Shuanghuanglian possibly can decrease Glu level and increase GABA content by way of its main component baicalin to antagonize the aconitine neurotoxicity.

OBJECTIVE:To study the effect and its mechanism of Fuzheng Jiedu Quyu formula(short forFuzheng formula) combined with oxaliplatin (L-OHP) on human colon cancer HT-29 cell proliferation and apoptosis. METHODS:HT-29 cells were divided into blank control group(without drugs),Fuzheng formula group(1000 mg/L),L-OHP group(31.25 mg/L)and combi-nation group (1000 mg/L Fuzheng formula+31.25 mg/L L-OHP). After cultured with corresponding drug for 48 h,MTT method was used to detect the cell proliferation;the changes of cellular morphology were observed by invert microscope;flow cytometry was used to detect the cell cycle and apoptosis rate. Proapoptotic gene Bax,apoptotic gene Bcl-2 mRNA expressions were deter-mined by real-time fluorescence quantitative polymerase chain reaction method;Bax,Bcl-2 protein expressions were assayed by Western blot. RESULTS:Compared with blank control group,cell proliferation was inhibited in L-OHP group and combination group;cell proportion was increased in S stage,G2/M stage and decreased in G0/G1 stage(P<0.05). Cell apoptosis rate in L-OHP group,Fuzheng formula group and combination group was increased;Bax mRNA and protein expression were up-regulated,Bcl-2 mRNA expression was downregulated(P<0.05);and combination group changed more obviously than the single drug groups(P<0.05). CONCLUSIONS:Fuzheng formula combined with L-OHP can inhibit HT-29 cell proliferation and promote its apoptosis, showing better effects than either of the two drugs alone. The mechanism may be associated with up-regulation of Bax gene and pro-tein expressions and down-regulation of Bcl-2 gene expressions in cells.