Chronic administration of growth hormone-releasing factor increases wound strength and collagen maturation in granulation tissue

The effects of chronic administration of growth hormone-releasing factor (GRF) on wound healing were studied in rats. Cutaneous wound strength was measured by tensometry at 5, 10, and 14 days postwounding in rats implanted with a slow-release pellet which contained a compressed mixture of a fatty acid and [desamino Tyr1, D-Ala2, Ala15]hGRF(1-29)NH2 or the fatty acid alone. There was a significant increase in wound tensile strength in GRF-treated rats compared to controls at each measurement: Day 5, 130 +/- 12 vs 97 +/- 14 g; Day 10, 402 +/- 18 vs 280 +/- 11 g; Day 14, 830 +/- 17 vs 614 +/- 14 g (P less than 0.01 for each value). Granulation tissue obtained from subcutaneously implanted polyvinyl alcohol sponges encased in silicone tubing was also studied. The amount of collagen deposited in the granulation tissue was estimated by measuring the hydroxyproline (Hyp) content of sponges retrieved 5, 10, and 14 days postinsertion from GRF-treated and control rats. Hyp content (nmole/mg sponge) was similar in both treated and control animals at each measurement: Day 5, 1.7 +/- 0.2 vs 2.2 +/- 0.2; Day 10, 31.9 +/- 4.1 vs 26.7 + 0.4; and Day 14, 41.6 +/- 7.3 vs 38.5 +/- 4.4. Hyp/proline, Hyp/glycine, and glycine/total amino acid ratios, evaluated after 10 days, were also similar in both groups. Collagen from the granulation tissue of sponges retrieved after 14 days from treated and control rats was studied by electron microscopy (magnifications, 7,100 and 22,720).(ABSTRACT TRUNCATED AT 250 WORDS)     

Treatment of Human Pancreatic Cancer in Mice with Angiogenic Inhibitors

Tumor growth is dependent on the balance of positive and negative regulators of angiogenesis. Antiangiogenic compounds inhibit endothelial cell biology in vitro and angiogenesis in vivo. Therefore antiangiogenic therapy presumes to be an effective treatment for pancreatic cancer. We wanted to determine the effect of antiangiogenic therapy on the growth of human pancreatic cancer in a mouse model. The angiogenesis inhibitors TNP-470 and antiangiogenic antithrombin III (aaATIII) were tested in vitro for their ability to inhibit endothelial cell proliferation. These inhibitors, along with the known antiangiogenic molecule endostatin, were then employed to treat two different primary human pancreatic cancers implanted subcutaneously into the dorsa of immunodeficient (SCID) mice. Treated tumors were examined histologically for microvessel density, apoptosis, and proliferation. All three inhibitors suppressed the growth of pancreatic tumors in vivo. Immunohistochemical analysis revealed increased degrees of apoptosis and reduced microvessel density in treated tumors compared to untreated tumors, although tumor cell proliferation was the same in both groups. None of the inhibitors tested significantly inhibited proliferation of human pancreatic cancer cells, although both TNP-470 and aaATIII were able to inhibit the proliferation of endothelial cells. The observed tumor suppression may be due to increased tumor cell apoptosis as a result of capillary dropout. These studies show that after the angiogenic switch in a human tumor, there is residual production of angiogenesis inhibitors.     

Effect of a smooth muscle antagonist on contraction of patched intestinal defects.

Growing new intestinal mucosa on serosal patches may potentially increase intestinal surface area but is limited by contraction of the serosal patch. Since this might be related to smooth muscle contraction or altered collagen metabolism, our aim was to determine whether the smooth muscle antagonist thiphenamil inhibits contraction. Fifty rabbits had two 2 x 5-cm full-thickness intestinal defects patched with adjacent cecum. Group I (n = 25) received saline and Group II (n = 25) 0.02 M thiphenamil at 10 cc/hr intraluminally. Animals were sacrificed at 1, 3, 5, 7, and 10 days. Group II had significantly less contraction of the proximal patch until the 10th day after patching (84 +/- 8 vs 66 +/- 20% Day 1, 67 +/- 4 vs 52 +/- 9% Day 5, 42 +/- 14 vs 42 +/- 7% Day 10). Epithelialization of patches was significantly less in Group II animals at 10 days (88 +/- 8 and 86 +/- 11% vs 47 +/- 20 and 50 +/- 16%, P less than 0.05) but crypt cell production rate and villus height were similar. The hydroxyproline content of regenerating tissue increased significantly 7 and 10 days after patching but was similar in the two groups (30.8 +/- 5.9 micrograms/mg tissue Day 10 vs 12.8 +/- 2.8 Day 1). Smooth muscle antagonism by thiphenamil inhibited contraction of serosal patches but had a deleterious effect on epithelialization and mucosal enzyme activity. The transient effect of thiphenamil and the associated increase of hydroxyproline content suggest that collagen may have the predominant role in contraction.     

Epidermal growth factor increases granulation tissue formation dose dependently

Recent studies have shown that epidermal growth factor (EGF) stimulated the rate of formation of granulation tissue in a model of wound repair (A. Buckley, et al., Proc. Nat. Acad. Sci. USA 82: 7340, 1985). Because pharmacologic doses of EGF were used previously, the relationship of EGF concentration to physiologic effects was determined in this study. Rats were implanted with subcutaneous polyvinyl alcohol sponges containing slow-release pellets formulated to release 0, 0.1, 1.0, or 10 micrograms of EGF/day. Tissue response was judged by the degree of histologic organization and vascularity, as well as several quantitative parameters: wet weight, hydroxyproline content, protein content, and DNA concentration. Each of these parameters showed consistent increases by Day 5 after implantation, when inflammation and edema had subsided. Compared with placebo controls, hydroxyproline (collagen) content was significantly increased by as little as 1 microgram/day of EGF, and DNA content was significantly increased by all dose levels of EGF. Endogenous EGF concentration in experimental granulation tissue was found to be fairly constant (30-40 ng/g wet wt); however, the increasing cellularity of the sponges may have reduced the local concentration of free EGF to low levels. Pellets releasing as little as 4 ng/hr of EGF into the surrounding tissue were able to accelerate wound healing, suggesting that the availability of this growth factor may be a rate-limiting step in wound repair.     

Troponin i peptide (Glu94-Leu123), a cartilage-derived angiogenesis inhibitor: In vitro and in vivo effects on human endothelial cells and on pancreatic cancer

Several inhibitors of angiogenesis have been identified in bovine and shark cartilage. One of them is troponin I, which is the molecule responsible for the inhibition of the actomyosin ATPase during muscle contraction. In this study we sought to investigate if the active site of troponin I (peptide Glu94-Leu123; pTnI) is also the one responsible for the antiangiogenic properties of this protein. The effects of pTnI on endothelial cell tube formation and endothelial cell division were investigated using human umbilical vein endothelial cells, Matrigel, light microscopy, carboxyfluorescein diacetate, succinimidyl esterlabeling, and flow cytometry. Its effects on induction of ICAM-1 and production of vascular endothelial growth factor by pancreatic cancer cells (CAPAN-1) were also investigated, as was its efficacy in a mouse model of pancreatic cancer metastases. Our results show that concentrations as low as 1 pg/ml of pTnI significantly inhibit endothelial cell tube formation, and that endothelial cell division was inhibited at 96 hours by 3 μg/ml pTnI (P = 0.0001). No effects were seen using troponin peptide 124-181 as a control. pTnI-treated supernatant from the pancreatic cancer cell line CAPAN-1 downregulated ICAM-1 expression on human umbilical vein endothelial cells up to 10 ng/ml pTnI, and a significant reduction in vascular endothelial growth factor production was seen by treating CAPAN-1 cells with up to 1 μg/ ml pTnI. After intrasplenic injection of CAPAN-1 cells, mice treated with pTnI had fewer liver metastases compared to control mice (liver/body weight 5.5 vs. 11.1; P = 0.03). The active region of troponin I is the one responsible for its antiangiogenic effect. The mechanism of action of this peptide is probably multifactorial. Presented at the Forty-Forth Annual Meeting of The Society for Surgery of the Alimentary Tract, Orlando, Florida, May 17–22, 2003 (oral presentation). Dr. Kern is supported by the Roche Research Foundation, Basel, Switzerland; the Novartis Stiftung, Basel, Switzerland; and the St. Clarastiftung, Basel, Switzerland.     

Wound healing: captopril, an angiogenesis inhibitor, and Staphylococcus aureus peptidoglycan

BACKGROUND: Captopril, an angiotensin-converting enzyme inhibitor, used for treating hypertension and heart failure, inhibits angiogenesis in the corneas of rats in response to basic fibroblast growth factor, slows the growth of experimental tumors in rats, and leads to the regression of Kaposi's sarcoma. Because angiogenesis is key to wound healing, we hypothesized that captopril would impair wound healing. We hypothesized also that because local application at operation of Staphylococcus aureus peptidoglycan (SaPG) increases angiogenesis and accelerates wound healing in rats, SaPG would prevent or ameliorate the postulated captopril-impaired wound healing. MATERIALS AND METHODS: In each experiment, rats were divided randomly into two groups: one drinking tap water, and the other, tap water containing 0.5 mg captopril/ml. All ate chow and drank ad libitum, pre-operatively (4-12 days) and postoperatively (7 days). In experiments 1 and 2, bilateral paravertebral 5.5-cm skin incisions were made aseptically (intraperitoneal sodium pentobarbital), and closed with interrupted No. 35 stainless-steel sutures. On one side, the wound was immediately inoculated with 157 microliter pyrogen-free isotonic saline and on the other side the wound was inoculated with 157 microliter saline containing 4.7 mg SaPG (860 microgram SaPG/cm incision). In the third experiment, polyvinyl alcohol (PVA) sponges (16-17 mg dry wt each) containing either 50 microliter saline or 0.5 mg SaPG in 50 microliter saline were implanted subcutaneously, two on each side, via 1-cm incisions closed with a single suture. In the fourth experiment, 5.5-cm bilateral skin incisions and subcutaneous implantation of PVA sponges were done as described but all sites were instilled with saline only. All rats were euthanized (CO(2) asphyxia) 7 days postoperatively. RESULTS: Wound breaking strength (WBS) of the saline-treated incisions was significantly higher (P < 0.001) in captopril-treated rats than in controls (172 +/- 13 g vs 105 +/- 6 g) in experiment 1 and higher, but not significantly in captopril-treated rats in experiment 2 (153 +/- 8 g vs 114 +/- 6 g) (PNS). SaPG inoculation of the incisions increased WBS significantly in both control and captopril-treated rats: 187 +/- 11 g vs 105 +/- 6 g (P < 0.001) and 283 +/- 16 g vs 172 +/- 13 g (P < 0.001), respectively, in experiment 1, and 217 +/- 13 g vs 114 +/- 6 g (P < 0.0001) (controls) and 266 +/- 17 g vs 153 +/- 8 g (captopril-treated rats) (P < 0.0001) in experiment 2. In experiment 3, subcutaneous PVA saline-inoculated sponge reparative tissue hydroxyproline (OHP) content was similar in control and captopril-treated rats, and SaPG inoculation increased reparative tissue OHP significantly in both groups: 2458 +/- 218 microgram/100 mg dry sponge vs 3869 +/- 230 microgram/100 mg (P < 0.001) (controls) and 2489 +/- 166 microgram/100 mg vs 4176 +/- 418 microgram/100 mg (P < 0.001) (captopril-treated rats). Histologically, angiogenesis and reparative tissue collagen were similar in control and captopril-treated rats, in both saline-inoculated and SaPG-inoculated sponges. In experiment 4 (all incisions and subcutaneous PVA sponges were saline-inoculated), there was no significant difference in WBS between control and captopril-treated rats (107 +/- 6 g vs 96 +/- 5 g, NS). PVA sponge reparative tissue OHP was significantly higher in captopril-treated rats: 3698 +/- 170 microgram/100 mg dry sponge vs 2534 +/- 100 microgram/100 mg (P < 0.0001). CONCLUSION: Unexpectedly, in four experiments, captopril did not inhibit WBS or PVA sponge reparative tissue angiogenesis or collagen accumulation; in fact, WBS was increased significantly in one of three experiments, and PVA sponge reparative tissue OHP was increased significantly in one of two experiments. Also, captopril did not interfere with the wound healing-accelerating effect of SaPG.     

鲨鱼软骨制剂辅助治疗晚期胆道恶性肿瘤的初步临床观察

目的：评价鲨鱼软骨制剂辅助治疗胆道晚期肿瘤的临床应用价值。方法：以寿命表法统计分析２９例应用鲨鱼软骨制剂的胆道恶性肿瘤患者与对照组的生存率。结果：鲨鱼软骨制剂治疗组的平均生存时间为２６个月,而对照组仅为８．５个月,治疗组明显长与对照组。结论：鲨鱼软骨制剂在辅助治疗晚期胆道恶性肿瘤中有一定的临床应用价值。     

Increased fibrovascular invasion of subcutaneous polyvinyl alcohol sponges in SPARC-null mice

The expression of SPARC (secreted protein acidic and rich in cysteine/osteonectin/BM-40) is elevated in endothelial cells participating in angiogenesis in vitro and in vivo. SPARC acts on endothelial cells to elicit changes in cell shape and to inhibit cell cycle progression. In addition, SPARC binds to and diminishes the mitotic activity of vascular endothelial growth factor. To determine the effect(s) of SPARC on angiogenic responses in vivo, we implanted polyvinyl alcohol sponges subcutaneously into wild-type and SPARC-null mice. On days 12 and 20 following implantation, SPARC-null mice showed increased cellular invasion of the sponges in comparison to wild-type mice. Areas of the sponge with the highest cell density exhibited the highest numbers of vascular profiles in both wild-type and SPARC-null animals. The endothelial component of the vessels was substantiated by immunoreactivity with three different markers specific for endothelial cells. Although sponges from SPARC-null relative to wild-type mice were populated by significantly more cells and blood vessels, an increase in the ratio of vascular to nonvascular cells was not apparent. No differences in the percentage of proliferating cells within the sponge were detected between wild-type and SPARC-null sections. However, elevated levels of vascular endothelial growth factor were associated with sponges from SPARC-null versus wild-type mice. An increase in vascular endothelial growth factor production was also observed in SPARC-null primary dermal fibroblasts relative to those of wild-type cells. In conclusion, we have shown that the fibrovascular invasion of polyvinyl alcohol sponges is enhanced in mice lacking SPARC, and we propose that increased levels of vascular endothelial growth factor account, at least in part, for this response.     

VEGF-mediated angiogenesis of human pheochromocytomas is associated to malignancy and inhibited by anti-VEGF antibodies in experimental tumors

BACKGROUND: Pheochromocytomas are well-vascularized tumors of the adrenal medulla. In human pheochromocytomas, angiogenesis has been associated with tumor progression. The mechanisms, however, are unknown. METHODS: Surgical specimens of benign, invasive, and metastatic human pheochromocytomas (n = 10/5/5) were immunostained for vascular endothelial growth factor (VEGF) and CD34, to determine VEGF expression and microvessel density (vascular surface density, [VSD]). In PC12-pheochromocytoma cells, VEGF messenger RNA and protein were analyzed by Northern blotting and enzyme immunoassay; biologic activity by human umbilical vein endothelial cell-proliferation assay. Inhibition of angiogenesis of PC12 xenografts by 2 neutralizing anti-VEGF antibodies (C20-pAB, M461-mAB) was evaluated by VEGF expression, VSD, and mitotic activity. RESULTS: VEGF expression and VSD were significantly higher in metastatic pheochromocytomas (VEGF 37.1 +/- 10.9% vs 20.7 +/- 9%, VSD 26.2 +/- 8 vs 13.5 +/- 3.3 1/mm). VEGF messenger RNA and protein were confirmed in PC12 cells and stimulated by nerve growth factor. Conditioned PC12 medium increased human umbilical vein endothelial cell proliferation more than 2-fold. Xentrotransplanted PC12 cells had marked VEGF expression and angiogenesis, which was inhibited by anti-VEGF antibodies (VEGF-expression by 29 and 38%, VSD by 43 and 46%, P <.05). CONCLUSION: Higher VEGF expression and microvessel density in malignant pheochromocytomas suggest VEGF-mediated angiogenesis to be related to tumor progression. Angiogenesis of PC12 xenografts is mediated by VEGF. Neutralizing anti-VEGF antibodies inhibit angiogenesis in experimental pheochromocytomas and may have potential for treating nonresectable pheochromocytomas.     

Longterm recombinant adeno-associated, virus-mediated, liver-generated expression of an angiogenesis inhibitor improves survival in mice with disseminated neuroblastoma

BACKGROUND: A gene therapy-mediated approach to the delivery of antiangiogenic agents using adeno-associated virus (AAV) vectors has a number of advantages including the potential for sustained expression. The purpose of this study was to attempt to restrict the growth of disseminated neuroblastoma through delivery of a truncated, soluble form of the vascular endothelial growth factor receptor-2 (VEGFR-2 fetal liver kinase [Flk]-1), a decoy receptor for VEGF. STUDY DESIGN: Mice underwent portal vein injection of vector, either AAV-CAGG-tsFlk-1 or control vector. Subsequent truncated soluble fetal liver kinase-1 (tsFlk-1) expression was confirmed and quantified by ELISA. After 8 weeks, mice were given human neuroblastoma cells through the tail vein to establish disseminated disease and were sacrificed after 40 days. The weight of liver metastasis was measured. Intrahepatic tumor microvessel density and levels of apoptosis were determined. Survival was assessed in a second cohort of mice. RESULTS: After intraportal injection of AAV-CAGG-tsFlk-1, high-level, stable transgene expression was generated. Sera from these mice inhibited endothelial cell activation in vitro and Matrigel plug (BD Biosciences) neovascularization in vivo, suggesting that a systemic state of angiogenesis inhibition had been established. After neuroblastoma injection, mice expressing tsFlk-1 had a smaller tumor burden in the liver when sacrificed, as compared with control mice. The decrease in tumor weight in the liver correlated with lower intratumoral microvessel density and higher levels of tumor cell apoptosis in the tsFlk-1-treated tumors, supporting the hypothesis that decreased angiogenesis had occurred. In a second cohort of mice, survival of tsFlk-1-expressing mice after tumor cell challenge was longer than in control mice. CONCLUSIONS: Longterm in vivo expression of a functional VEGF inhibitor was established using an AAV-mediated gene therapy approach, and it demonstrated antiangiogenic and anticancer efficacy in a murine model of disseminated neuroblastoma.     

Wound healing in nude mice: a study on the regulatory role of lymphocytes in fibroplasia

In order to understand the role of T cells in postinjury fibroplasia, we have studied wound healing in congenitally athymic nude mice that lack a normally developed T cell system. Healing of incisional wounds, as assessed by wound breaking strength, was significantly stronger in nude mice compared with normal thymus-bearing animals. This was accompanied by a marked increase in the amount of reparative collagen synthesized at the wound site, as assessed by the hydroxyproline content of subcutaneously implanted sponges. Because nude mice have some extrathymic T cell maturation, we used an anti-Thy-1.2 (30H12) monoclonal antibody to selectively deplete T cells in vivo. Although such treatments impaired wound healing in normal mice, they had no effect on any wound healing parameter in nude mice. In a separate experiment, T cell reconstitution of nude mice, sufficient to significantly enhance in vivo delayed hypersensitivity responses, led to a decrease in both wound breaking strength and hydroxyproline deposition in subcutaneously implanted polyvinyl sponges. The data suggest that T cells play a dual role in wound healing: an early stimulatory role on macrophages, endothelial cells, and fibroblasts, and a late counterregulatory role, which may be responsible for the orderly completion of wound repair.     

Effect of the intraperitoneal paclitaxel on the healing of colonic anastomoses

The effect of the antiangiogenic agent paclitaxel on the healing of colonic anastomoses was studied in a rat model. Ninety-six rats underwent end-to-end colonic anastomoses, and healing was evaluated by measuring bursting pressure, hydroxyproline content, and number of newly formed vessels. The rats with end-to-end anastomoses were randomized into control and study groups (n = 48). The rats in the control group were administrated 5 ml physiologic saline intraperitoneally, and those in the study group were given 4.5 mg/kg paclitaxel in 5 ml physiologic saline intraperitoneally. One-half of the rats in both groups were killed on day 4, and the other half were killed on day 7 after surgery with high doses of ether anesthesia. There were no deaths. No detectable improvement in anastomoses bursting pressure was observed in the paclitaxel group on day 4 after surgery. On postoperative day 7, a statistically significant difference was found between the groups (P < 0.05). On day 4 after surgery, there was no statistically significant difference in hydroxyproline levels and vessel density between the two groups (P > 0.05; hydroxyproline: 26.25 versus 16 microg/mg tissue; vessel contents: 110.8 versus 118.2 vessels/x 100 fields for control and study groups, respectively). On postoperative day 7, a significant difference was detected between the two groups (P < 0.0001; hydroxyproline: 40.5 versus 9.6 microg/mg tissue; P < 0.05; vessel contents: 105 versus 85.3 vessels/x 100 fields for control and study groups, respectively). Neovascularization and hydroxyproline levels in rats receiving paclitaxel were significantly reduced on postoperative day 7 (P < 0.05 and P < 0.0001, respectively). These findings suggest that the antiangiogenic efficiency of paclitaxel, and thus, negative impact on wound healing, is more prominent on postoperative day 7. Experimentally, paclitaxel may inhibit and delay healing of colonic anastomoses.     

Effect of rapamycin on wound healing: an experimental study

The aim of this study was to investigate the effects of rapamycin (RAPA) on the healing of bladder and abdominal wound closures. Fourteen male Sprague Dawley rats were randomized to receive either RAPA (3 mg/d) or placebo. A midline laparotomy was performed. The bladder was cut and closed with 4-0 Vicryl in a double layer. The fascia was closed with 0 nylon suture, and the skin closed with a subcuticular 2-0 nylon suture. The mean RAPA level was 9.1 ng/mg. Eosinophil and neutrophil infiltration, and the presence and degree of myofibroblast proliferation were significantly higher in the bladder, fascia, and dermis of the control group. Lymphocyte infiltration was similar in each group. Mean microvessel density as well as the percentage of cells expressing vascular endothelial growth factor in the bladder, fascia, and dermis were significantly lower among the RAPA group. Both proliferating cell nuclear antigen labeling indices for inflammatory cells in the fascia, dermal fibroblasts, and epithelial cells in the placebo group were significantly higher. No difference was observed for hydroxyproline levels in both the bladder and fascia between the groups. In conclusion, we found that RAPA treatment affected all steps of the wound healing process by decreasing the inflammatory cell number, angiogenesis, and myofibroblast proliferation, so the wound healing process was delayed and consequently the tensile strength of the wound decreased.     

Amelioration of microvascular myocardial ischemia by gene transfer of vascular endothelial growth factor in rabbits

OBJECTIVES: Restoration of coronary blood flow by angiogenesis may offer a new approach to intractable ischemic heart disease. In the present study, we investigated the angiogenic effects of gene transfer of vascular endothelial growth factor 165 on microvascular myocardial ischemia. METHODS: A rabbit model of microvascular myocardial ischemia was created by plugging coronary microvessels with microspheres (15 microm in diameter, 2.8 x 10(5)/kg, n = 29). Gene transfer was performed by semi-selective intracoronary injection of recombinant adenovirus expressing vascular endothelial growth factor 165 forty minutes after microsphere injection (n = 9). RESULTS: Microsphere injection reduced myocardial perfusion (78% +/- 9% of baseline tissue flow) and diminished myocardial contraction (61% +/- 12% of the baseline ejection fraction) and cardiac performance (elevated left ventricular end-diastolic pressure and decreased systemic flow) in the acute phase. At 17 +/- 3 days, gene transfer of vascular endothelial growth factor 165 had had the following effects: (1) promoted coronary angiogenesis as evidenced by myocardial flow above the baseline (121% +/- 24%), (2) increased vascular density revealed by synchrotron radiation microangiography and histologic analysis, (3) ameliorated the degree of myocardial ischemia as evidenced by myocardial lactate content and the extent of histologic necrosis, and (4) restored heart function as evidenced by increased ejection fraction (95% +/- 10%), reduced left ventricular end-diastolic pressure, and restored body weight. CONCLUSIONS: In vivo vascular endothelial growth factor 165 gene transfer promoted angiogenesis and was an effective approach to treating microvascular myocardial ischemia.     

Antitumor and antiangiogenic effects of somatostatin receptor-targeted in situ radiation with (111)In-DTPA-JIC 2DL

INTRODUCTION: Expression of somatostatin receptor subtype 2 (sst 2) in angiogenic tumor vessels appears to be homogeneous, while tumor cell expression of this receptor is often heterogeneous. We have developed a novel in vitro three-dimensional tumor angiogenesis model to study the antitumor and the antiangiogenic effects of radiolabeled somatostatin analogs. We hypothesized that targeted in situ radiation with an Auger electron-emitting radiolabeled somatostatin analog would produce receptor-specific cytotoxicity in sst 2-expressing cells. MATERIALS and METHODS: IMR-32 human neuroblastoma (sst 2-positive) and MDA MB-231 human breast cancer (sst 2-negative) xenografts were created in nude mice from monolayer cell cultures. Fragments of these tumors were embedded in three-dimensional fibrin gels supplemented with endothelial growth media and incubated for a period of 14 days. Tumor fragments were treated with 50 microCi/ml of (111)In-JIC 2DL, a sst 2-preferring somatostatin analog, or medium on Day 1. Initial angiogenic activity was determined at 48 h and the mean angiogenic score and tumoricidal responses were assessed on Day 14. RESULTS and CONCLUSION: Tumoricidal effects of (111)In-JIC 2DL were seen only in sst 2-positive IMR-32 tumors. However, the angiogenic response was inhibited in both IMR-32 and MDA MB-231 tumors independent of the tumor cells' sst 2 status. Somatostatin receptor-mediated in situ radiation therapy has profound cytotoxic effects on angiogenic blood vessels and sst 2-expressing tumor cells.     

兔缺血后肢内源性VEGF蛋白表达与侧支形成相关性研究

目的：探索缺血肢体代偿侧支形成过程中内源性ＶＥＧＦ蛋白表达的相应规律。方法：新西兰雄性白兔４０只,切除兔一侧后肢股浅动脉全长建立缺血模型,分为１天、３天、７天、１４天、２８天、５６天及９０天７个观察期（ｎ＝５）;另设对照组（ｎ＝５）。观察造模后小腿动脉压比率、动脉造影血管计数及缺血后肢肌组织毛细血管密度和内源性ＶＥＧＦ蛋白的变化。结果：造模后３天,缺血后肢骨骼肌中即有内源性ＶＥＧＦ蛋白表达,７天时达到高峰,２８天后逐渐减弱,而缺血区新血管发生也相应停滞。结论：内源性ＶＥＧＦ蛋白表达有其特定时限性,提示在其表达水平开始下降时,应考虑补充外源性ＶＥＧＦ。     

大鼠失神经靶肌肉注射异体不同细胞对神经再生影响的研究

目的研究注射异体不同细胞于大鼠失神经靶肌肉对神经再生的影响.方法 SD成年大鼠36只,雌性,体重120～150 g,随机分为4组,每组9只.无菌条件下切断左侧坐骨神经,一期神经外膜缝合.术后即于小腿三头肌处注射相应细胞,7天注射1次,共4次.A组注射单纯雪旺细胞1×106/ml 1 ml,B组注射雪旺细胞加成肌细胞(雪旺细胞∶成肌细胞为1∶1)1×106/ml 1 ml,C组注射肾内皮细胞提取液1 ml,D组注射无血清培养液1 ml作对照.术后3个月取手术侧坐骨神经及坐骨神经所支配的小腿三头肌,进行大体和组织形态学观察、神经鞘细胞密度及单位面积靶肌肉(小腿三头肌)运动终板计数.结果术后3个月,各组近端神经鞘细胞均数为A组0.134 5±0.029 8,B组0.093 1±0.025 6,C组0.072 4±0.023 7,D组0.187 7±0.054 2;A组∶D组P值<0.05,B组∶D组、C组∶D组及A组∶B组P值均<0.01,B组∶C组P值＞0.05.术后3个月各组远端神经鞘细胞密度均数为A组0.186 0±0.042 5,B组0.155 1±0.032 1,C组0.104 7±0.013 3,D组0.240 9±0.056 8;A组∶D组P值<0.05,B组∶D组、C组∶D组及B组∶C组P值<0.01,A组∶B组P值＞0.05.术后3个月各组靶肌肉运动终板个数均数为A组6.000±0.866,B组9.000±2.291,C组12.780±1.394,D组3.110±0.782;A组∶D组、B组∶D组及C组∶D组P值<0.01,A组∶B组、A组∶C组及B组∶C组P值<0.01.结论雪旺细胞、混合细胞、肾内皮细胞提取液均有促进神经再生作用,肾内皮细胞提取液优于雪旺细胞及混合细胞.     

Experimental study of intraperitoneal suramin on the healing of colonic anastomoses.

INTRODUCTION: The effect of the antiangiogenic agent suramin on the healing of colonic anastomoses was studied in a rat model. METHODS: Rats underwent an end-to-end colonic anastomosis and its healing was tested by measuring bursting pressure, hydroxyproline content and number of newly formed vessels. For the bursting pressure experiment suramin was given intraperitoneally in a dose of 200 mg/kg (maximal tolerable dose) and 100 mg/kg. Hydroxyproline content and vessel density were only tested at 100 mg/kg since the toxicity at this dose was lower whereas bursting pressure was still diminished. RESULTS: There were no deaths. On the fourth day after operation bursting pressure in the control group was significantly higher than that in rats treated with suramin 200 mg/kg (P = 0.006) and 100 mg/kg (P = 0.002). Rupture occurred at the anastomotic line. On day 7, this difference was not statistically significant. Four days after the operation, the hydroxyproline content and vessel density were significantly below that in control segments (hydroxyproline: 10.3 versus 7.8 microg per mg dry weight; vessel content: 85.7 versus 49.6 vessels per x 100 field for control and suramin-treated rats respectively). On the seventh day no difference in hydroxyproline levels was seen but the vessel density was still diminished significantly (P = 0.04). CONCLUSION: Experimentally, suramin significantly inhibits and delays healing of colonic anastomoses. Presented in part to the XXXII Congress of the European Society for Surgical Research in Corfu, Greece, May 1997     

成人骨髓间充质干细胞分化为血管内皮细胞的研究

目的研究成人骨髓间充质干细胞（marrow mesenchymal stem cells，MSCs）体外分化为血管内皮细胞的能力并探讨其诱导条件。方法分离成人骨髓单个核细胞，经贴壁法获取MSCs。所获细胞分4组进行诱导：组1、2以5×10^3／cm^2细胞密度接分别接种于含2％和15％胎牛血清的L—DMEM培养基；组3、4以5×10^4／cm^2细胞密度分别接种于上述两种血清浓度的培养基，各组细胞经血管内皮生长因子（vascular endothelial growth factor，VEGF）辅以牛脑提取物的诱导，对照组为正常传代培养的MSCs。在诱导的第14、21天分别测定表面分子CD34、VEGFR-2、CD31和Ⅶ因子相关抗原（factor Ⅷ—related antigen，又称von Willebrand factor，vWF）以及体外成血管实验对分化细胞进行鉴定，同时测定不同诱导条件下的细胞增殖指数（proliferation index，PI）。结果组3细胞经诱导后，内皮系表面分子CD34、VEGFR-2、CD31、vWF于第14天均有不同程度表达，分别为8．5％、12．0％、40．0％、30．0％．其中CD34、VEGFR-2在第21天表达上调，为15．5％、20．0％，余各诱导组细胞未表达上述表面分子；诱导后的组3细胞呈低增殖状态（PI值约为10．4％），在半固体培养基中还可形成血管腔样结构。结论从成人骨髓中分离培养的MSCs，在较高细胞接种密度和低增殖状态下，经VEGF、牛脑垂体提取物诱导后，可分化为血管内皮细胞，可作为治疗性血管生成及组织工程理想的种子细胞。     

Angiogenesis by endothelial cell transplantation.

PURPOSE: Myocardial angiogenesis may improve regional perfusion and perhaps function after cardiac injury. We evaluated the effect of endothelial cell transplantation into a myocardial scar on angiogenesis and ventricular function, as an alternative to angiogenic gene or protein therapy.Methods and Results: A transmural myocardial scar was created in the left ventricular free wall of rat hearts by cryoinjury. Allogeneic aortic endothelial cells were injected into the scar 2 weeks after cryoinjury. A cluster of transplanted cells was identified at the site of injection 1 day and 1 week after transplantation, but not after 2 weeks. The size of this cluster of transplanted cells decreased as vascular density in the transplanted scar tissue increased with time. Six weeks after transplantation, vascular density was significantly greater in transplanted hearts than in control hearts. Regional blood flow, by microsphere analysis, was greater in the transplanted rats. Systolic and diastolic ventricular function was similar between groups. In a second series of experiments, syngeneic aortic endothelial cells labeled with bromodeoxyuridine were transplanted 2 weeks after cryoinjury. Vascular density in the transplanted scar was greater than in controls. Labeled transplanted endothelial cells were identified forming part of the newly developed blood vessels. No difference in vascular density was found between allogeneic and syngeneic cell transplantation. Vascular endothelial growth factor was not expressed at greater levels in the transplanted cells or the myocardial scar. CONCLUSION: Transplanted endothelial cells stimulated angiogenesis, were incorporated into the new vessels, and increased regional perfusion in myocardial scar tissue, but did not improve global function in this cryoinjury rat model.