BACKGROUND: The islet cell transplantation has provided a solid basis for diabetic therapy, but the insufficient donor limits its development.OBJECTIVE: improving the method of isolating and purifying islets to observe the transplantation effect.DESIGN: A laboratory animal research.SETTING: Key Laboratory of Animal and Department of Cell Biology, China Medical University.MATERIALS: The experiment was carried out in the Key Laboratory of Animal and Department of Cell Biology in China Medical University between January and October in 2006. Donors were Wistar rats of either gender, weight 250-300 g;Acceptors were SD male rats, weight 180-220g. The two kinds of rats were all common closed population and from the Experimental Animal Department of China Medical University (The Admission Number of Experimental Animal Institute is SYXK(LIAO)2003-0013).METHODS: ①Isolation and exaltation of islet cells as well as the functional evaluation of pancreas: After etherisation, the Wistar rat without fasting was executed. A little cut was made on the beginning of the biliary pore, then the little cut lumbar anesthesia ductus, which were connected with a 1-mm-diameter syringe and full of cold collagenase solution (1.5 g/L), was inserted directly to dilate pancreas thoroughly. The pancreatic gland was isolated and digested in the water of centrifuge, when doing that, 1 mol/L NaOH was put interruptedly into the centrifuge tube to keep the pH value of the solution at 7.8±1.0. The rat pancreas purified by centrifugation of Ficoll density gradient: The identification of purified islets was evaluated by dithizone staining. The viability of islet was assessed by fluorescence staining of aridine orange and propidium iodide. The motility rate=the total number of live cells/(the total number of live cells + the total number of dead cells)×100%. Pancreatic activity was calculated: insulin release index=the level of insulin at the third hour (high concentration glucose)/the level of insulin at the second hour (low concentration glucose). ②The blood from vena caudalis of SD rat was sampled and measured the blood sugar after the intraperitoneal injection of streptozocin. The rat was diagnosed as DM when blood sugar was more than 16.7 mmol/L twice without fasting. The DM rats were divided into two groups, every group 8 rats. The experimental group rats were injected about 1 000 islet cells into the location below renal capsule, and the control group rats were injected the same volume of 1640 cultu re solution. Eight normal rats, whose glucose concentration ≤ 5.5 mmol/L, were taken randomly as normal controlled group. The blood sugar was measured every day after the surgery. The blood sugar less than 11.1 mmol/L without fasting was taken as the sign of successful islet transplantation. Intravenous sugar tolerance test was applied to the rats of normal control group, DM control group and experimental group 3 days after islet transplantation. Fasting for 12 hours before test, the blood sugar was measured at 0, 15, 30, 60, 90 and 120 minutes.MAIN OUTCOME MEASURES: Purity quotient, survival rate and activity of islet cells.RESULTS: All 24 SD acceptor rats were involved in the result analysis without miss.①The total number of purified islets of one pancreas was (1 150±141) in well morphology. The purity of islets was more than 95%. The viability of islets was more than 98%. ②The insulin secretion response to glucose challenge in vitro showed the mean value of insulin in the low-glucose medium was (70.5±6.9) mlU/L, while that of high-glucose medium was (321.4±11.6) mlU/L, the insulin release index was 4.6±0.52, that meant the beta cell of islet functioned well. ③The blood glucose level and the insulin level in plasma of the transplanted recipients restored to normal 3 days after transplantation. The survival period of transplanted islets was (6±2) days. But there was not any change in the concentration of blood sugar in the control group (16.7 mmol/L). The intravenous glucose tolerance test showed the identical outcome between the islet splantation group and the normal control group.CONCLUSION: There are high yield and high purify of islet cells in rats, which are isolated by in situ perfusion and purified by Ficoll density gradient centrifugation.

Objective: The inhibition on the proliferation of colorectal carcinoma cell strain with different proliferative potential by ATRA was investigated in present study, which would benefit for the therapy of ATRA on colorectal carcinoma. Methods: The proliferation of LS174T and CW-2 colorectal carcinoma cell strain inhibited by ATRA was determined using cell observation, FACS and MTT methods. Results: The growth speed of LS174T cell strain was faster than that of CW-2. ATRA played a significant role on inhibiting the proliferation of LS174T and CW-2 cell strain and inducting the cell differetiation in vitro. Conclusions: ATRA could inhibit the growth of LS174T and CW-2 cell strain. ATRA could inhibit the proliferation of colorectal carcinoma cell and induce cell differentiation to some extent, which was correlated with the concentration of ATRA.

Objective To investigate the influence of menstrucal cycle and anatomic site on the fractional anisotropy (FA) values of diffusion tensor imaging (DTI) in normal breast. Methods Prospectively enrolled 96 volunteers, who have identified with normal menstrucal phase and without breast diseases were found via the breast examination, ultrasound and MRI scan. The cases were divided into three groups according to menstrucal phase: menstrual period group(menstrual cramps 1 to 6 d), proliferative phase group(menstrual cramps 7 to 14 d) and secretory phase group(menstrual cramps 15 d to the next), and each group consisted of 32 subjects. All subjects were performed bilateral breast cross-sectional T1WI, T2WI, DWI and DTI scaning. On the nipple level figture, the mammary gland was divided into three regions including the anterior, central and posterior parts, and the FA values of the different phases and regions were measured. The Kruskal-Wallis H test was applied to analyse the difference of FA values in different menstrual phase and anatomic site. Results The FA values of the anterior region in menstrual phase, proliferative phase and secretary phase were 0.21 ± 0.07, 0.24 ± 0.09 and 0.17 ± 0.07, and the difference had significant difference(P=0.014).The FA values of the central region were respectively 0.15±0.08, 0.18±0.09 and 0.15±0.07, and without the statistically significant difference(P=0.090). The FA values of the posterior region were 0.21 ± 0.11, 0.24 ± 0.13 and 0.16 ± 0.11, and also showed significant difference(P=0.002). In different regions, the difference of FA values between menstrual phases and proliferative phases were also had statistically significant(P=0.018, 0.045, respectively). In the same region, the FA value was lowest in the secretary phase, and the proliferative phase was slightly higher than menstrual phase. Conclusion The FA values are affected by menstrual cycle and anatomic site.

Objective:To study the effect of early hyperbaric oxygen combined with rehabilitation training on neurological rehabilitation and prognosis of patients with hypertensive intracerebral hemorrhage after borehole drainage.Methods:Eighty-five patients with hypertensive intracerebral hemorrhage admitted to Zhejiang Xin'an International Hospital from January 2018 to March 2020 were enrolled, and all of them were treated with minimally invasive drilling and drainage and they were divided into two groups by the order of admission. The control group (41 cases) received rehabilitation training after 48 h of stable vital signs. The research group(44 cases) was treated on the base of the control group combined with early hyperbaric oxygen therapy. The levels of neuron-specific enolase (NSE), S100β and scores of National Institutes of Health Stroke Scale (NIHSS), Fugl-Meyer Assessment (FMA), Barthel index, cerebral blood flow, and Glasgow Outcome Scale (GOS) were observed in the two groups before and after treatment.Results:After treatment, the levels of NSE and S100β in the two groups were decreased and those in the research group were lower than those in the control group: (14.38 ± 1.47) μg/L vs. (18.04 ± 2.95) μg/L, (0.24 ± 0.03) μg/L vs. (0.32 ± 0.04) μg/L, the differences were statistically significant ( P<0.05). The scores of NIHSS in the two groups were decreased, while the scores of FMA and Barthel index were increased, while the scores of NIHSS in the research group was lower than that in the control group: (9.18 ± 1.02) scores vs. (11.85 ± 1.47) scores; the scores of FMA and Barthel index in the research group were higher than those in the control group: (68.38 ± 8.93) scores vs.(61.42 ± 8.01) scores, (63.25 ± 7.65) scores vs. (56.11 ± 6.04) scores, the differences were statistically significant ( P<0.05). The average cerebrovascular blood flow and flow rate of patients in both groups were higher than those before treatment, and the peripheral cerebrovascular resistance of patients in both groups was lower than that before treatment, while the average cerebrovascular blood flow and flow rate in the research group were higher than those in the control group: (17.85 ± 2.48) ml/s vs. (15.12 ± 2.01) ml/s, (20.61 ± 2.88) cm/s vs. (17.04 ± 2.47) cm/s; the peripheral cerebrovascular resistance in the research group were lower than that in the control group: (1 328.95 ± 114.29) kPa·s/m vs. (1 558.13 ± 157.21)kPa·s/m, the differences were statistically significant ( P<0.05). The good prognosis rate of research group was higher than that of control group: 84.09%(37/44) vs. 60.98%(25/41), the difference was statistically significant ( χ2 = 4.634, P<0.05). Conclusions:Early hyperbaric oxygen combined with rehabilitation training can promote the recovery of neurological function of patients with hypertensive intracerebral hemorrhage after borehole drainage, and can improve cerebral blood flow and prognosis.

BACKGROUND: To apply mouse anti-human cTnI monoclonal antibody as the drug vector in the treatment and diagnosis of myocardial injury, it is important to degrade the immunity of murine antibody and overcome human anti-mouse reaction. Humanization has been applied as an attempt to resolve this problem.OBJECTIVE: To clone murine anti-cTnI Fab fragment and analyse the nucleotide and deduced amino acid sequences.DESIGN: Single sample study.SETTING: An institute of cardiovascular disease under a medical university-affiliated hospitalMATERIALS: The study was conducted in the Institute of Cardiovascular Diseases, First Affiliated Hospital of Nanjing Medical University from January 2003 to May 2004. The hybridoma cell line JS200202 which secrets the anti-cTnI monoclonal antibody was provided by Institute of Cardiovascular Disease, First Affiliated Hospital of Nanjing Medical University.METHODS: IgG heavy chain primers and κ light chain primers of amplified mouse were designed. Total RNA was extracted from hybridoma cells which secrete cTnI. Reverse transcription polymerase chain reaction(RT-PCR) was amplified. Cloning and subsequent sequence analysis of the Fab fragment was performed. The deduced amino acid sequence was compared and analysed with previously published sequences.MAIN OUTCOME MEASURES: Heavy chain Fd segment and κ light chain gene sequence and its subgroups.RESULTS: A band of approximate 700 and 800 base pairs were amplified using IgG heavy chain primers and κ light chain primers respectively. Sequence analysis indicated that the deduced amino acid sequences were in consistent with the characterization of the amino acid in the murine IgGl Fab fragment(GenBank accession NO AY484430, AY484431; Protein Bank accession NO AAR83243, AAR83244).CONCLUSION: A complete murine anti-cTnI Fab fragment was obtained in this study, which may provide basis for the production of the chimeric anti-cTnI antibody.

Objective To explore the application of continuous subcutaneous insulin infusion (CSII) in elderly type 2 diabetic patients. Methods Elderly group(n = 415) and control group (n= 461) received the transient CSII intensive treatment. The different regiments and the incidence of hypoglycemia were observed between the two groups. Results The average duration reaching the target blood glucose and the insulin dosage at the target time were similar between the two groups (P>0.05). The insulin basal rate in the elderly patients was lower than that in control group. The incidence of hypoglycemia, especially at night, was significantly higher in the elderly patients.Conclusions CSII could control blood glucose effectively in elderly type 2 diabetic patients, but the basal insulin dosage should be decreased to reduce the risk of hypoglycemia.

Objective: To study expressions of small ubiquitin-related modifier protein（SUMO）4 (SUMO4), nuclear factor (NF)- κB and inhibitory factor of NF-κB (IκB) in kidneys of rats with type 2 diabetes mellitus (T2DM). Methods: A total of ten 40-week-old male Goto-Kakizaki (GK) rats （with spontaneous diabetes mellitus）of specific-pathogen free (SPF) grade, and ten 40-week-old male Wistar rats of SPF grade were selected. The lesion of renal tissue was observed by hematoxylin eosin (HE) staining. Expressions of SUMO4, NF-κB and IκB in renal tissue were observed by immunohistochemistry methods. Results: In the GK rats, glomerular capillary ball hypertrophy, basilar membrane slightly thickening; glomerular mesangial cells hyperplasia, hypertrophy and renal tubular epithelial cells hypertrophy were observed. Compared with normal Wistar rats, expression levels of NF-κB [(0.232±0.034) vs. (0.634±0.058)], IκB [(0.242±0.027) vs. (0.712±0.078)] and SUMO4 [(0.160±0.031) vs. (0.545±0.045)] significantly increased in renal tissue of GK rats (P<0.01 all). Conclusion: Compared with Wistar rats, expressions of NF-κB, IκB and SUMO4 significantly increase in renal tissue of GK rats, suggesting that SUMO inhibiting transcriptional activity of NF-κB may exist in kidneys of T2DM rats. Therefore, sumoylation may be a new therapeutic target for inhibit renal microvascular lesion of diabetic disease.

Objective: To study relative risk factors for diabetic nephropathy (DN). Methods: A total of 238 patients diagnosed as type 2 diabetes mellitus (DM) were enrolled in the study. According to urine microalbuminuria to urine creatinine ratio (UACR), patients were divided into pure DM group (group DM1, n=90), early diabetic nephropathy group (group DM2 , n=73) and clinical diabetic nephropathy group (group DM3 ,n=75). Clinic data of all patients were collected; Fasting blood glucose (FBG), 2h postprandial blood glucose (2hPB), blood lipids, uric acid (UA), fibrinogen (Fg) and glycosylated haemoglobin (HbA1c) were measured in all patients, and their correlations with DN were analyzed. Results: Compared with group DM1, the course of disease in DM [(7.25±6.29) years vs. (10.25±7.67) years vs. (13.53±7.82) years], levels of FBG [(8.46±2.52) mmol/L vs. (9.52±3.38) mmol/L vs. (10.82±3.30) mmol/L], 2hPB [(18.40±5.64) mmol/L vs. (20.27±5.94) mmol/L vs. (22.59±6.14) mmol/L], HbA1c [(7.96±1.65) % vs. (8.60±1.76) % vs. (9.55±2.09) %], triglyceride [TG, (1.72±0.86) mmol/L vs. (2.34±1.87) mmol/L vs. (3.16±1.85) mmol/L], Fg [(3.49±0.93) g/L vs. (3.88±1.21) g/L vs. (4.99±2.10) g/L] and UA [(295.42±52.34) μmol/L vs. (324.18±96.29) μmol/L vs. (351.23±56.88) μmol/L] significantly increased in group DM2 and group DM3 in order (P<0.01~0.001). Logistic gradual regression analysis indicated that course of DM, HbA1c, TG, Fg and UA were risk factors for DN (OR=1.008~1.910, P<0.01~0.001). Conclusion: The course of DM, blood glucose, blood lipid, uric acid and fibrinogen are risk factors of diabetic nephropathy; increased UACR reflects progress of patient’ condition in DM patients, its detection is used for diabetic prognosis and treatment.

The blood-brain barrier (BBB) prevents most therapeutic drugs from entering the brain tissue, which makes diagnosis and treatment of brain diseases difficult, especially diseases of central nervous system. Focused ultrasound can open BBB non-invasively and repeatedly by means of microbubble contrast agents. In recent years, experimental and clinical researches on focused ultrasound treatment of brain diseases have been carried out at home and abroad. The research progresses of focused ultrasound in brain diseases were reviewed in this article.

Objective To explore the effect of adiponectin on the pathophysiology and treatment of type 2 diabetes. Methods 50 patients with newly diagnosed type 2 diabetes were given insulin intensive therapy.Body mass index(BMI), blood lipids, blood glucose, insulin and adiponectin were measured before and after the insulin treatment.36 healthy persons were selected to match for sex and age with diabetes patients. Results In type 2 diabetes patients, before insulin intensive treatment,the level of adiponcetin was 4.28(2.11-6.38)μg/L and arose to 12.26(8.22-16.00)μg/L after 12.6±2.1 day insulin treatment.In healthy persons, the level of adiponectin was 9.56(7.92-11.61)μg/L.There was significant difference among the three groups.Adiponectin was negatively correlated with BMI and triglyceride and positively correlated with insulin action index (IAI) and high density lipoprotein (HDL). Conclusions Adiponectin plays a major role in the pathophysiology and treatment of newly diagnosed type 2 diabetes.