BACKGROUND:Intervertebral disc degeneration is the pathological basis of degenerative spinal diseases. Studies on the influentialfactors of intervertebral disc degeneration contribute to the prevention and treatment of degenerative spinal disease.
OBJECTIVE:To observe the growth and proliferation of nucleus pulposus cels isolated by trypsin plus type II colagenase digestion in complete medium with different glucose concentrations, exploring the optimal glucose concentration for growth of nucleus pulposus cels.
METHODS:Nucleus pulposus cels isolated and cultured by trypsin plus type II colagenase digestion method were observed under an inverted microscope, and thecelnumber was counted. Morphology of nucleus pulposus cels was observed afterhematoxylin-eosinstaining and toluidine blue staining. Colagen type II immunoreactivity was detected by immunohistochemical staining combined with immunofluorescent staining.Nucleus pulposuscels were incubated in complete medium containing various glucose concentrations (0, 6.25, 12.5, 17.5, and 25 mmol/L) for 24 hours, and then cel proliferation and apoptosis were determined.
RESULTS AND CONCLUSION:The stained nucleus pulposus cels showed polygonal and short spindle, with one or two nuclei. Celularpseudopod appeared gradualy and then became slim with increased passage numbers. The isolated and cultured nucleus pulposus cels positively expressed colagen type II and aggrecanProliferative activity of nucleus pulposus cels cultured in medium with 17.5 mmol/L glucose was significantly higher than that in medium with 0 and 25 mmol/L glucose (P< 0.05 orP< 0.01). There wasno significant differencein cel apoptosis between these groups except for 0 mmol/L glucose (P<0.05). These results confirm that a large number of nucleus pulposus celscan beharvested by trypsin plustype II colagenase digestion and the optimal glucose concentration is 17.5 mmol/L.

The drug resistant mutations in human immunodefieiency virus type 1 (HIV-1) are a major impediment to successful highly active antiretrovirai therapy (HAART) and new drug design. In order to understand the drug resistance mechanism of HIV-1 integrase (IN) mutually existed for multiple drug-resistant strains to the most potent IN inhibitors diketo acids (DKAs), three S-1360-resistant HIV-1 strains were selected and molecular docking and molecular dynamics (MD) simulations were performed to obtain the inhibitor binding modes. Based on the binding modes, compelling differences between the wild-type and the 3 mutants for IN have been observed. The results showed that: 1) In the mutants, the inhibitor is close to the funetional loop 3 region but far away from the DNA binding site. Different binding sites lead to the decrease in susceptibility to S-1360 in mutants compared to the wild-type IN. 2) The fluctuations in the region of residues 138～166 are important to the biological function of IN. 2 hydrogen-bonds between S-1360 with residues N155 and K159 restrict the flexibility of the region. Drug resistant mutations result in a lack of the interaction, consequently, the less susceptible to S-1360. 3) In the 3 mutant IN complexes, the benzyl ring of S-1360 is far from the viral DNA binding site, thus, S-1360 can not prevent the end of the viral DNA from exposure to human DNA. 4) After T66I mutation, the long side chain of I occupied the active pocket in the 3 mutants, consequently, the inhibitor could not move into the same binding site or have the same orientation. All the above contribute to drug resistance. These results will be useful for the rational inhibitor modify and design.

Objective To investigate the clinical effect and pathogenesis of tiopronin in treating acute pancreatitis complicated with liver damages. Methods Eighty-one patients with acute pancreatitis complicated with liver damages were randomly divided into two groups. The control group (40 cases) received routine treatment, and the treatment group (41 cases ) received routine treatment and tiopronin. The liver function, pancreatic enzyme indexes and C-reactive protein (CRP), the length of hospital stay was recorded. Results After 8 days' treatment, the liver function improved, and the levels of pancreatic enzyme indexes and CRP were decreased significanfly in both groups, and these changes were more obvious in treatment group (P <0.05). The length of hospital stay of mild acute pancreatitis in treatment group was significantly shorter than that in control group [ (8.6 ± 2.7 ) d vs. ( 13.8 ± 3.5 ) d ] (P < 0.05 ). Conclusions Tiopronin has beneficial effects in treating acute pancreatitis complicated with liver damages,and it has a therapeutic effect in acute pancreatitis at the same time. The mechanism may be related with the inhibition of inflammatory mediators, oxygen free radicals scavenging and other effects.

Objective:To assess the left ventricular (LV) systolic function in amateur marathoners by two-dimensional speckle tracking echocardiography.Methods:A total of 59 amateur marathon runners were recruited from January 2019 to June 2020 in Hangzhou and were divided into group A (>2-5 years) and group B (>5 years) based on their time of participating in marathon. Thirty-one healthy volunteers were enrolled from Affiliate Hospital of Hangzhou Normal University during the same period as a control group. Conventional echocardiography combined with two-dimensional speckle tracking imaging were applied to all the subjects to obtain interventricular septum diastolic thickness (IVSd), LV posterior wall thickness(PWd), LV end-diastolic diameter (LVEDd), relative wall thickness(RWT) and LV mass(LVM), LV end-diastolic volume(EDV), LV end-systolic volume(ESV) and stroke volume(SV), LV ejection fraction(LVEF), LV global longitudinal strain (GLS), and global circumferential strain (GCS). Pre-marathon, 1 hour and 4th day post-marathon echocardiography were performed in amateur marathon runners.Results:Compared with the control group, group A amateur marathon runners showed significant increases in IVSd, PWd, LVEDd, RWT and LVM(all P<0.05); In addition, the IVSd, PWd, LVEDd, RWT, LVM, EDV, ESV and SV in group B runners were further increased compared to those of group A runners (all P<0.01). The LVEF and pre-marathon GCS showed no statistically significant differences among the three groups (all P>0.05), while the pre-marathon GLS showed a statistically significant difference among the three groups ( P<0.01). Compared with group A runners, the pre-marathon, 1 hour and 4th day post-marathon GLS and 1 hour post-marathon GCS were significantly decreased in group B runners (all P<0.01); In intra-group comparison, 1 hour post-marathon GLS was significantly decreased in relative to pre-marathon and 4th day post-marathon GLS in both group A and B (all P<0.05). One hour post-marathon GCS was significantly decreased compared with pre-marathon GCS in group B ( P<0.05). Conclusions:Amateur runners who have participated in long time marathon have reduced LV longitudinal strains and transient post-marathon decreases in LV systolic function.

Objective To investigate the effectiveness of simple peritoneal lavage combined with venous-venous hemofiltration therapy for severe acute pancreatitis (SAP). Methods Needles were inserted into abdominal cavity and tee was connected, then normal saline was administrated and discharged, followed by lidocaine, dexamethasone and antibiotics once daily until bloody peritoneal drainage became clear. At the same time venous-venous hemofiltration was used. Results 61 SAP patients were randomly divided into peritoneal lavage + hemofiltration group (treatment group, n =31) and control group (n = 31). The time to abdominal pain relief, abdominal distention relief, nausea and vomiting disappearance, peritoneal irritation disappearance was (1.5 ±0.3)d,(2.7 ±0.3)d, (1.9 ±0.3)d, (1.5 ±0.2)d, and the time to cure was (11.0 ±2.0)d in the treatment group, which was significantly shorter than that in the control group [(3.9 ± 0. 3) d, (4.5 ±0.6)d, (3.7 ±0.2)d, (5.3 ±0.4)d, (18.0 ±2.5)d, P＜0.05]. At the 1st day of treatment, serum ALT,AST was significantly lower than that in the control group; at the 3rd day of treatment, the serum and urine amylase and serum levels of TNF-α, IL-6 and IL-8 level were significantly lower, but the serum level of IL-10,HCO3-was significantly higher than that in the control group (P ＜ 0.05 or ＜ 0. 01); at the 5th day of treatment, the serum Bun and Cr level were significantly lower than those in the control group (P ＜ 0. 05).Conclusions Simple peritoneal lavage combined with venous-venous hemofiltration therapy can effectively eliminate the inflammatory factors, which is more rational and effective for the treatment of severe acute pancreatitis.

ObjectiveTo study changes of molecular markers of prothrombotic state:Platelet granule membrane protein ( GMP-140 ),Von Willebrand factor ( vWF:Ag),thrombomodulin (TM),Two-D dimer ( DD),antithrombin Ⅲ ( AT- Ⅲ ) in plasma and puerarin for treatment functions of acute pancreatitis (AP).MethodsIn 78 patients with AP [ severe acute pancreatitis (SAP):26 cases,mild acute pancreatitis (MAP):52 cases ],using a random number table,the patients were given puerarin treated base (n =40) and conventional treated base group (n =38 ).The two groups were given fast,continuous gastrointestinal decompression,correction of electrolyte and acid-base balance disorders,vein support,antisecretory drugs,antibiotics inhibit pancreatic secretion and inhibition of trypsin activity of drug treatment.Puerarin group:Puerarin injection 0.5 g in 5％glucose injection intravenous infusion of 500 ml,1 time a day.GMP-140 vWF:Ag,TM,DD were measured by the methods of analysis of enzyme-linked immunosorbent assay and AT-Ⅲ was measured by the methods of analysis of chromogenic substrate method preformed in all patients,plasma amylase and uric amylase were determined by the method of somogyi and after the treatment.And 22 healthy people were selected as normal controls ( NC,Group C,n =22).ResultsCompared with the Group C and MAP,the plasma GMP-140 [ ( 86.26 ± 15.28 )ng/Lvs (32.56 ± 18.17) ng/L and (58.68 ± 15.86)ng/L],vWF[(236.22 ±31.78)％vs (95.12 ±31.68)％ and (126.68 ± 17.06)％ ],TM [(65.70 ± 12.27) μg/L vs (4.26 ±0.92) μg/L and (9.80 ± 6.98) μg,/L],DD [ (0.87 ±0.04) mg/L vs (0.36 ±0.06) mg/L and (0.56 ±0.05) mg/L] were significantly elevated,however the AT-Ⅲ [ (56.13 ± 15.78) U/ml vs (98.76 ±22.68) U/ml and (80.38 ± 18.29)U/ml )was significantly decreased SAP ( P ＜ 0.01 ).There were significant differences on the levels of GMP-140 [ (31.52 ± 15.81 ) ng/L vs (59.62 ± 13.73 ) ng/L,t =- 23.283 ],vWF [ ( 93.32 ± 28.62) ％ vs ( 128.81 ±16.23)％,t=-28.205,P＜0.01 ],TM[ (4.36 ± 0.82) μg,/L vs (11.23 ± 7.62)μg/L,t =-43.419,P ＜0.001],DD[ (0.32 ±0.05) mg/L vs (0.68 ±0.04) mg/L,t =- 15.642,P ＜0.001],AT-Ⅲ ((97.68 ±21.69) U/ml vs (76.86 ± 17.92) U/m,t =14.967,P ＜ 0.01 ) between puerarin treated base group and conventional treated base group.Comparing with treated base,the group given puerarin obviously shortened the increased of plasma [ ( 81.26 ± 17.12) U/L vs ( 119.63 ± 51.87 ) U/L,t =- 7.618,P ＜ 0.001 ],uric amylase [ (416.37 ± 116.50) U/L vs (576.32 ± 126.58) U/L,t =- 36.659,P ＜ 0.001 ],the time of abdominal pain relief and therapy to spend [ ( 2.18 ± 0.76 ) d vs ( 5.26 ± 0.58 ) d,t =- 13.619,P ＜ 0.001 ].Conclusion The molecular markers of prothrombotic state:GMP-140,vWF:Ag,TM,DD,AT- Ⅲ might all play key roles in the development of AP.Puerarin can improve the pancreatic microcirculation and adjust molecular markers of prothrombotic state,and had certain treatment functions with AP.

ObjectiveTo study the changes of the plasma thrombomodulin(TM) and von Willebrand factor (vWF) levels and their clinical significance associated with the extent and severity of acute ischemie colitis.MethodsThe plasma TM and vWF levels were determined by enzyme linked immunosorbent assay in 46 patients with acute ischemic colitis (acute ischemic colitis group),42 patients with ulcerative colitis (ulcerative colitis group) and 40 healthy subjects (control group).ResultsThe plasma TM was (49.6 ±2.3) μg/L,and vWF was(198.8 ±8.9)％ in acute ischemic colitis group.The plasma TM was (38.2 ± 3.8) μ g/L,and vWF was ( 162.6 ± 7.6)％ in ulcerative colitis group.The plasma TM was (23.8 ±2.3) μg/L,and vWF was ( 116.7 ± 6.2)％ in control group.The plasma TM and vWF levels in acute ischemic colitis group were higher than those in ulcerative colitis group and control group (P ＜ 0.05 or ＜ 0.01 ).The plasma TM and vWF levels in ulcerative colitis group were higher than those in control group (P＜ 0.05).The plasma TM levels[(49.9 ± 0.3 ) μg/L] and vWF [(210.6 ± 8.2 ) ％] in all colon disease were higher than those in partial colon disease (P ＜ 0.05 ).ConclusionThe changes of plasma TM and vWF levels can be used as one of the indicators for assessment of the development and the prognosis of acute ischemic colitis.

Objective To study changes of endothelin (ET),nitric oxide (NO) in plasma and treatment effect of puerarin on acute pancreatitis (AP). Methods Seventy-three patients with AP were randomly divided into 2 groups: puerarin group (38 cases, puerarin plus basic treatment) and non puerarin group (35 cases, basic treatment ). ET was measured by radioimmunoassay, NO was measured by nitrate reductase and plasma amylase and uric amylase were determined by Somogyi before and after the treatment.Twenty healthy persons were recruited as control group. Results The levels of ET[(40.6±15.8) ng/L]and NO[(62.3±27.6) mmol/L] decreased after treatment in puerarin group, which were significantly lower than those in non puerarin group[(82.3±20.6) ng/L, (92.6±24.8 ) mmol/L]( P < 0.05 ). Compared with those in non puerarin group[(1182.0±520.0), (5623.0±1326.0) U/L and (5.12±0.76)d], plasma amylase and uric amylase [(802.0±170.0), (4102.0±1126.0) U/L], the time of abdominal pain relief [(2.20±0.72) d] was lower in puerarin group (P<0.05). Conclusions The plasma ET and NO might play a key role in the development of AP. Therapeutic effect of puerarin is sure in AP.

BACKGROUND:Intervertebral disc degeneration is caused by damage and degeneration of the nucleus pulposus and annulus fibrosus tissues inside the intervertebral disc,resulting in structural and functional changes of the intervertebral disc.However,there is yet no effective drug treatment for intervertebral disc degeneration. OBJECTIVE:To investigate the inhibitory effect of syringin on intervertebral disc degeneration. METHODS:A total of 10 male Sprague-Dawley rats were selected,and the coccygeal intervertebral disc(Co4/Co5)of each rat was set as model group,Co5/Co6 intervertebral disc as syringin group,and Co6/Co7 intervertebral disc as control group.The control group did not receive any treatment.In the model group and syringin group,a miniature puncture needle was used to puncture the annulus fibrosus to establish an intervertebral disc degeneration model.Immediately after modeling,2.5 μL of normal saline and syringin solution(5 μmol/L)were given in the model and syringin groups,respectively.Four weeks after injection,the samples were taken.The degree of intervertebral disc degeneration in rats was observed by hematoxylin-eosin and safranine O-fast green staining.The expressions of type Ⅱ collagen,aggrecan and matrix metalloproteinases 3 and 13 in intervertebral disc tissue were analyzed by immunohistochemical staining. RESULTS AND CONCLUSION:Hematoxylin-eosin staining showed that in the model group,the height of intervertebral disc decreased,the cartilage endplate became thinner and cracked,the fibrous ring structure was disordered and cracked,and the nucleus pulposus disappeared;in the syringin group,the height of intervertebral disc was normal or slightly lower than that in the control group,the degree of cartilage endplate degeneration was lighter than that in the model group,the fiber circle permutation was relatively regular with no cracks,and the nucleus pulposus was partially shrunk.Safranine O-fast green staining showed that in the model group,the cartilage endplate of the intervertebral disc was defective and the calcified layer of cartilage became thinner,showing obvious degeneration.The structure and morphology of intervertebral disc cartilage endplate in the syringin group recovered to some extent.Immunohistochemical staining showed that,compared with the control group,the expressions of type Ⅱ collagen and aggrecan in the intervertebral disc cartilage were decreased in the model group(P<0.000 1),while the expressions of matrix metalloproteinases 3 and 13 increased(P<0.000 1).Compared with the model group,the expressions of type Ⅱ collagen and aggrecan in the intervertebral disc cartilage tissue were increased in the syringin group(P<0.001,P<0.000 1),while the expressions of matrix metalloproteinases 3 and 13 decreased(P<0.001,P<0.000 1).These results showed that syringin could improve the structure and function of intervertebral disc by inhibiting the expression of matrix metalloproteinases 3 and 13 and increasing the expression of type Ⅱ collagen and aggrecan,thus preventing and slowing down the procession of intervertebral disc degeneration.

Objective:To assess the left ventricular(LV) structure and systolic function in amateur marathon runners using real-time three-dimensional speckle tracking echocardiography(3D-STE) and analyze its correlation with the running volume.Methods:A total of 84 amateur marathon runners were recruited between January 2019 and October 2021 in Hangzhou and were divided into short-term (ST) group(≤6 months) and more extended-term(MET) group(>6 months–2 years) based on their time of participating in the marathon. Thirty-nine healthy volunteers were enrolled from the Affiliated Hospital of Hangzhou Normal University during the same period as a control group. The running volume of ST and MET runners were recorded, LV end-diastolic volume(EDV), end-systolic volume(ESV), LV mass(LVM), LV ejection fraction(LVEF), LV global longitudinal strain(GLS), global circumferential strain(GCS), global radial strain(GRS), global area strain (GAS), twist, and torsion were measured by conventional echocardiography and 3D-STE. The differences of those parameters among the three groups were compared. The correlation between 3D-STE parameters and the running volume was further analyzed.Results:In ST amateur marathon runners, LV EDV was higher compared with controls( P<0.05). Compared with ST and control groups, LV EDV, ESV, and LVM of MET runners were increased(all P<0.05). In addition, compared with control group, GLS of MET runners was increased( P<0.05). The LVEF, GCS, GRS, GAS, twist, and torsion showed no statistically significant differences among the three groups(all P>0.05). In amateur marathon runners LV EDV, ESV, LVM had statistically significant positive correlations with an average weekly running volume and total running volume (all P<0.01). Multivariate linear regression analysis showed that the total running volume was an independent correlation factor in LV EDV of amateur marathon runners (β=0.618, P<0.01). Conclusions:Amateur marathon runners participating in a short term marathon mainly show an increase in LV EDV, the longitudinal systolic function of the LV can be enhanced in the early stage of the marathon.