A One–Step, Operator–Independent Method for Isolating Islets of Langerhans from the Porcine Pancreas

Abstract: Large–scale isolation of islets of Langerhans is one of the major obstacles in islet transplantation. Until now, isolation methods relied on enzymatic digestion, the duration of which relies on a decision dictated by the operator's experience. This approach has always hindered development of an automated method. The aim of this study was to develop a one–step method based on complete digestion of the pancreas. The original aspect of the technique (derived from the Ricordi method) is use of the University of Wisconsin (UW) solution in the digestion medium and a continuous flow collagenase processing circuit with local cooling and rewarming to allow tissue digestion to proceed at 37°C while settling of the cell suspension takes place at 4°C. A stopcock system permits the alternate use of two settling chambers so that while one is in the circuit, the other can be removed for cen–trifugation, resuspension of the crude islet preparation in collagenase in free UW solution, and further purification in a density gradient system. Ten experiments were performed, and 545, 750 ± 48, 670 purified pig islets were obtained per totally digested pancreas. Histological studies showed cell integrity. Insulin secretion in response to double glucose stimulation under perfusion conditions demonstrated the functional viability of the isolated islets. In conclusion, this one–step method makes it possible to obtain a high number of viable islets of Langerhans in the absence of any decision by an operator, and it should therefore provide basis for an automated method.     

Small Cell Carcinoma of the Prostate: A Case Report

A case of small cell carcinoma of the prostate without a primary lesion in the lung was reported. The cancer was diagnosed after the patient complained of lumbago caused by bone metastasis. The tumor was 5.9 times 5.0 times 4.6cm. The patient was treated with 4 courses of chemotherapy using cisplatin and etoposide. The tumor diminished to 4.0 times 4.0 times 3.5 cm after completion of the 4 courses of treatment. Prostatic antigen levels were less than 1.0ng/mL during the therapy. Neuron-specific enolase levels were 35.9ng/mL at the beginning of therapy, and decreased to 7.4 ng/mL after completion of 4 courses of treatment. The patient died 3 months after the completion of treatment. This regimen had some value for inhibiting the growth of small cell carcinoma.     

Enhanced Intestinal Absorption of Oxytocin Peptide Analogues in the Absence of Pancreatic Juice in Pigs

Purpose. The present investigation was done to study the intestinal absorption of three oxytocin peptide analogues and to elucidate the role of pancreatic juice on their absorption. Methods. In conscious chronically catheterized pigs (6–8 weeks of age) plasma concentration of the peptides, [Mpa¹, D-Tyr(Ethyl)², Thr⁴, Orn⁸]-oxytocin (F314), [Mpa¹, D-Tyr(Ethyl)², Val⁴, D-Arg⁸]-oxytocin (CAT), and [Mpa¹, D-Tyr(Ethyl)², Thr⁴, Orn⁸, desGly⁹, carba⁶]-oxytocin (F327) after intraduodenal administration, during presence or diversion of the pancreatic juice via a pancreatic duct catheter, were determined by radioimmunoassay. The stability of the peptides to degradation was determined in vitro by incubation with activated pancreatic juice, chymotrypsin or trypsin, followed by reversed phase HPLC analyses. Results. All peptides were absorbed with a bioavailability of about 0.5% in the presence of pancreatic juice, but increased to 1.0%, 2.1%, and 13.5% for F314, CAT, and F327, respectively, when the pancreatic juice was diverted from the intestine. After incubation with pancreatic juice 95% of F314, 98% of F327, and 100% of CAT was found intact. When incubated with trypsin CAT remained intact while F314 and F327 were degraded by 54% and 46%, respectively. Incubation with purified chymotrypsin did not degrade the test peptides. Conclusions. The results indicate that the increased absorption of peptides observed under conditions of diverted pancreatic juice cannot only be explained by the absence of pancreatic enzymes, but also by changed absorptive properties in the gastrointestinal tract.     

无毒棉籽液抗腹泻作用研究

通过对正常小鼠、生大黄致泻小鼠及脾虚小鼠的作用研究表明，无毒棉籽液有较好的抑制排便频度、治疗腹泻的作用．无毒棉籽液６ｇ／ｋｇ可极其显著地抑制正常小鼠的排便频度，无毒棉籽液６ｇ／ｋｇ和３ｇ／ｋｇ可极其显著地抑制生大黄致泻小鼠及脾虚小鼠的排便频度．６ｇ／ｋｇ无毒棉籽液且可极其显著地抑制正常小鼠及脾虚小鼠的小肠推进运动．     

新生儿肠旋转不良的诊治分析

目的 分析新生儿期先天性肠旋转不良的临床特点及诊治方法。方法 回顾分析1996～2003年12例新生儿期先天性肠旋转不良，男10例，女2例，就诊时出生7～75d，平均45d，术前均经泛影葡胺口服及灌肠造影确诊后行Ladd术，附加阑尾切除术。结果本组1例术后并发粘连性肠梗阻二次手术行粘连松解术，其余11例无并发症，全部治愈出院。结论 本病的发病机制是由于并发急性肠扭转所致高位十二指肠的梗阻，主要临床特点是反复间断性胆汁性呕吐，口服和灌肠的消化道造影是诊断本病的最可靠依据。Ladd式手术是本病手术治疗的最佳选择，且效果满意。     

Liver and intestine transplantation

The most significant development in liver transplantation in the USA over the past year was the full implementation of the MELD- and PELD-based allocation policy in March 2002, which shifted emphasis from waiting time within broad medical urgency status to prioritization by risk of waiting list death. The implementation of this system has led to a decrease in pretransplant mortality without increasing post-transplant mortality, despite a higher severity of illness at the time of transplant.
The trend over the last few years of rapidly increasing numbers of adult living donor liver transplants was reversed in 2002 by a decline of more than 30% in the number of these procedures. In 2002, a greater percentage of women received livers from living donors (43%) than deceased donors (34%), possibly because of size considerations.
From 1993 to 2001, the waiting list increased more than sixfold, from 2902 patients to 18 047 patients. For the first time since 1993, the waiting list size decreased in 2002, dropping 6% to 16 974 candidates. The percentage of temporarily inactive liver candidates also increased from 2001, thus the net decrease in the active waiting list for 2002 was 12%. This may reflect a trend toward less pre-emptive listing practices under MELD.
Intestine transplantation remains a low-volume procedure limited to a few transplant centers and is still accompanied by significant pre- and post-transplantation risks. As this procedure matures, its application may increase to include recipients at an earlier stage of their disease with better likelihood of success.     

Editorial

Aim: Wall stress‐independent signalling pathways were studied for endothelin‐1 (ET‐1)‐induced c‐fos expression in rat intact mesenteric small arteries. Methods: Arteries were kept unmounted in Krebs buffer, equilibrated for 1 h and stimulated with vasoactive substances for 15–60 min. The c‐fos mRNA expression was determined by real‐time polymerase chain reaction. Results: Stimulation with fetal bovine serum (FBS), phorbol 12‐myristate 13‐acetate (PMA) and ET‐1 caused about a doubling of c‐fos mRNA. The ET‐1‐induced c‐fos expression was steady (15–60 min) and was inhibited by the inhibitor of the ET_A receptor, BQ‐123. Platelet‐derived growth factor‐B, angiotensin II and U46619 did not cause increased c‐fos mRNA levels. The broad specificity inhibitor staurosporine inhibited the response to ET‐1, but inhibitors of Rho‐A kinase and phosphatidylinositol 3‐kinase had no effect. However, inhibitors to tyrosine kinases, the MAP kinases [extracellular signal‐regulated kinase 1/2 (ERK1/2), c‐Jun amino‐terminal kinase, p38], and to conventional protein kinase C showed no inhibition. Consistent with these findings, ET‐1 did not cause activation of ERK1/2, a finding also seen in vessels held under pressure. In contrast, ET‐1‐induced c‐fos expression was inhibited by the calcium chelator BAPTA, suggesting a role for intracellular calcium. This possibility was supported by the finding that raising the extracellular K⁺ concentration caused increased expression of c‐fos in a concentration‐dependent manner. Conclusion: The results suggest that in the absence of wall stress, ET‐1 is able to induce increased expression of c‐fos independent of traditional growth pathways, such as MAP kinase. The mechanism appears to be calcium‐dependent.     

非小细胞肺癌转移预测指标的研究

目的 :研究非小细胞肺癌 (NSCLC)淋巴结和远处转移的预测指标 ,并建立Logistic回归模型。　 方法 :通过免疫组化、ELISA、酶谱电泳等方法 ,对NSCLC肿瘤病理标本、血清、尿液和骨髓等进行检查 ,并通过Logistic回归分析建立预测概率模型。　结果 :免疫组化指标肿瘤组织内微血管密度 (IMVD)、血管内皮细胞生长因子(VEGF)、碱性成纤维细胞生长因子 (b FGF)、白细胞分化抗原变异型 (CD4 4v6 )、基质金属蛋白酶 2 (MMP 2 )与NSCLC淋巴结转移危险有关 (P <0 .0 5 ) ,组织金属蛋白酶抑制物 (TIMP 2 )、上皮型钙粘素 (E cad)与NSCLC淋巴结转移危险下降有关 (P <0 .0 5 )。血清MMP 2、MMP 9,尿液MMP 2、MMP 9及骨髓上皮膜抗原 (EMA)阳性细胞与NSCLC远处转移危险有关 (P <0 .0 5 )。其中免疫组化指标CD4 4v6、IMVD、E cad及尿液MMP 2、骨髓EMA阳性细胞对NSCLC转移有显著回归效果而分别被选入概率模型 1和 2 ,其预测准确率分别为 81.1%和 72 .7%。　结论 :组织标本中CD4 4v6、IMVD、E cad以及尿液中MMP 2及骨髓EMA阳性细胞检查 ,可预测绝大多数NSCLC的淋巴结转移和远处转移状况 ,为NSCLC转移的早期诊断提供重要信息 ,有助于NSCLC的个体化治疗和改善预后     

耕耘疗法在中小面积深度烧伤的临床应用

目的 :探讨耕耘疗法在中小面积深度烧伤创面治疗中的疗效。方法 :对小面积深Ⅱ度～浅Ⅲ度烧伤创面进行了耕耘疗法 ,然后外用MEBO。结果 :治愈 16 5例 ,深Ⅱ度创面多在 17± 2 0天内愈合。浅Ⅲ度创面也在 2 8± 3 0天内愈合。结论 :通过采用耕耘方法可改善深度创面的局部微循环 ,尽快清除坏死组织 ,促进创面液化 ,充分发挥MEBO药效 ,使创面愈合期明显缩短 ,减少了疤痕率     

大鼠小肠上皮细胞的体外原代培养

目的：探讨大鼠小肠上皮细胞原代培养的原理与方法以及培养过程中常见的问题和克服办法。方法：取生后1周SD大鼠的小肠上皮细胞，经分离纯化后进行原代培养。结果与结论：胶原酶Ⅺ型和中性蛋白酶Ⅰ型配合使用，可以分离到形态完整、容易贴壁生长的小肠绒毛细胞团。进一步利用离心技术纯化得到95％以上的小肠上皮细胞。结果显示，常规体外培养的肠上皮细胞是否增殖分化最终决定于用酶消化后的结构完整性，如果用0．25％胰酶消化，则细胞受损程度较大，不易贴壁生长。