兔胎仔腹裂模型实验研究

目的制作兔胎仔腹裂模型，探讨肠管病理损害机制和有关胎仔实验外科的技术。方法40只孕兔随机分成三组，分别于妊娠21d、23d、25d行宫内手术，每只孕兔制作胎仔模型1～3只，包括假手术、腹腔内注射羊水和腹裂三组，至孕30d时自然产仔或行剖宫产同收胎仔模型，分别测量出生体重、小肠重量、长度、密度、厚度，并观察肠壁的组织学和细胞超微结构变化。结果孕21d组腹裂模型存活率(37．04％)显著低于孕23d组(70．59％)和25d组(70．83％)；与对照组相比，腹裂模型组胎仔体重低，小肠重量轻、长度短、密度低、厚度增加，病理组织学和超微结构也发生改变。结论胎仔外科的成功受实验对象、胎龄、麻醉、宫内操作技术等诸多因素影响；腹裂胎仔肠管大体形态和组织结构的改变是受羊水浸泡和腹壁裂口缩窄共同作用的结果。     

乌司他丁对缺氧缺血性新生鼠的脑保护作用

目的 探讨乌司他丁对新生鼠缺氧缺血性脑损伤（HIBD）的作用。方法 选7日龄新生Wistar大鼠60只制备HIBD模型。乌司他丁治疗组大鼠分别于术后即刻、术后8、16h腹腔注射乌司他丁一次，术后24h处死。取脑组织迅速秤重，固定，切片行HE和TUNEL染色，测定凋亡细胞数，对皮层脑细胞变性坏死和胶质细胞增生行病理量化评分，电镜观察海马区细胞形态。结果 治疗组在脑重增加值及左、右脑重差值、凋亡细胞数和病理量化评分明显低于缺氧缺血组（P〈0．01）；各治疗组之间比较上述指标无统计学意义（P〉0．05）。结论 乌司他丁可减轻缺氧缺血后脑细胞水肿、变性、坏死及凋亡，对新生鼠HIBD具有保护作用。     

肠细胞凋亡在新生鼠坏死性小肠结肠炎肠损伤中动态变化

目的动态观察新生大鼠坏死性小肠结肠炎(NEC)发病过程中肠细胞凋亡率变化及其与肠损伤关系。方法40只新生SD大鼠随机分成对照组(C)和模型组(M)。对照组8只;模型组32只,在出生48h开始给予鼠配方奶人工喂养,100%氮气缺氧90s,4℃冷刺激10min,每天2次,连续3d,建立新生大鼠NEC模型;模型组开始造模后24h(M24)、48h(M48)、72h(造模结束,M72)及造模结束后24h(M96)分别处死8只,留取肠管进行肠组织损伤评分和肠细胞凋亡率检测(流式细胞仪)。组织学评分≥2确定为NEC。各组随机选取1份回盲部近端小肠标本进行肠黏膜透射电镜检查。采用SPSS11.0统计学软件进行统计分析,α=0.05为显著性检验标准。结果透射电镜显示模型组大鼠肠黏膜出现大量凋亡细胞,形成凋亡小体。对照组、M24、M48、M72和M96肠组织损伤评分分别为(0.08±0.15)、(1.38±0.42)、(1.46±0.69)、(1.58±0.30)分和(3.33±0.59)分,肠细胞凋亡率分别为4.8%±2.9%、12.8%±6.3%、14.9%±5.5%、17.7%±5.5%和27.6%±9.9%。肠损伤程度与肠细胞凋亡率呈显著正相关(r凋亡率=0.853,P<0.01)。结论新生鼠肠细胞凋亡增加是NEC肠组织损伤起始事件;随时间延长,肠细胞凋亡增加程度进一步加重;肠细胞凋亡增加是造成新生鼠NEC肠道进行性损伤的病理基础。     

肠细胞凋亡在新生鼠坏死性小肠结肠炎肠损伤中动态变化

目的 动态观察新生大鼠坏死性小肠结肠炎(NEC)发病过程中肠细胞凋亡率变化及其与肠损伤关系.方法 40只新生SD大鼠随机分成对照组(C)和模型组(M).对照组8只;模型组32只,在出生48 h开始给予鼠配方奶人工喂养,100%氮气缺氧90 s,4℃冷刺激10 min,每天2次,连续3 d,建立新生大鼠NEC模型;模型组开始造模后24 h(M24)、48 h(M48)、72 h(造模结束,M72)及造模结束后24 h(M96)分别处死8只,留取肠管进行肠组织损伤评分和肠细胞凋亡率检测(流式细胞仪).组织学评分≥2确定为NEC.各组随机选取1份回盲部近端小肠标本进行肠黏膜透射电镜检查.采用SPSS 11.0统计学软件进行统计分析,α =0.05为显著性检验标准.结果 透射电镜显示模型组大鼠肠黏膜出现大量凋亡细胞,形成凋亡小体.对照组、M24、M48、M72和M96肠组织损伤评分分别为(0.08±0.15)、(1.38±0.42)、(1.46±0.69)、(1.58±0.30)分和(3.33±0.59)分,肠细胞凋亡率分别为4.8%±2.9%、12.8%±6.3%、14.9%±5.5%、17.7%±5.5%和27.6%±9.9%.肠损伤程度与肠细胞凋亡率呈显著正相关(r＜凋亡率=0.853,P＜0.01).结论 新生鼠肠细胞凋亡增加是NEC肠组织损伤起始事件;随时间延长,肠细胞凋亡增加程度进一步加重;肠细胞凋亡增加是造成新生鼠NEC肠道进行性损伤的病理基础.     

先天性膈疝大鼠模型神经嵴源器官异常和心肺发育不良

目的 Nitrofen大鼠膈疝模型中的心血管畸形多是神经嵴源性的，我们由此推论，除了心肺发育不良外，同是神经嵴源性的甲状腺、甲状旁腺和胸腺也可能异常。本实验利用该模型研究其中神经嵴源器官的形态与生化改变，以及这些改变和膈疝之间的关系。方法 在怀孕第9．5d给予Sprague-Dawley母鼠100mg Nitrofen，第21d剖腹取出胎鼠。共使用16窝72只胎鼠(其中含对照组5窝20只)，随机分成形态组和生化组，研究其颈胸部内分泌腺体和心肺形态及心脏DNA和蛋白质／DNA.结果 发现Nitrofen处理后胎鼠常合并胸腺、甲状腺、甲状旁腺和心大血管崎形。心肺明显发育不良，膈疝的胎鼠尤其显著，而对照组全部正常。结论 该模型中心血管和肺的异常与临床所见一致，而胸颈部内分泌腺的异常值得我们注意。该模型是对CDH的病因、病理和合并畸形等方面研究的合适工具。     

乌司他丁对缺氧缺血性新生鼠的脑保护作用

目的探讨乌司他丁对新生鼠缺氧缺血性脑损伤(HIBD)的作用。方法选7日龄新生Wistar大鼠60只制备HIBD模型。乌司他丁治疗组大鼠分别于术后即刻、术后8、16h腹腔注射乌司他丁一次,术后24h处死。取脑组织迅速秤重,固定,切片行HE和TUNEL染色,测定凋亡细胞数,对皮层脑细胞变性坏死和胶质细胞增生行病理量化评分,电镜观察海马区细胞形态。结果治疗组在脑重增加值及左、右脑重差值、凋亡细胞数和病理量化评分明显低于缺氧缺血组(P<0·01);各治疗组之间比较上述指标无统计学意义(P>0·05)。结论乌司他丁可减轻缺氧缺血后脑细胞水肿、变性、坏死及凋亡,对新生鼠HIBD具有保护作用。     

先天性腹裂外科治疗的麻醉体会

目的总结新生儿先天性腹裂修补术的麻醉体会。方法回顾性分析23例新生儿先天性腹裂修补术的麻醉过程。结果3例采用全麻后Ⅰ期关腹直接复位缝合术，14例先采用非麻醉下免缝Silo袋Ⅰ期免缝手术，5～7d后在气管插管全麻下行Ⅱ期复位缝合术，只放置Silo袋、后期死亡或放弃治疗4例；未经任何治疗自动出院2例。手术患儿中12例麻醉过程较平稳，5例术中血压轻度下降，经加快输血输液等处理后血压回升，无麻醉死亡，均顺利完成手术。术后死亡3例，1例死于多脏器功能衰竭，2例术后肺膨胀不良死于呼吸衰竭。随访时间4个月至4年，患儿均生长发育正常。结论手术是治疗先天性腹裂的主要方法，新生儿先天性腹裂复位修补术的围手术处理关键是麻醉保障，而麻醉的重点是气道管理和保证循环功能稳定。     

叶酸缺乏孕鼠子代脑组织DNA甲基化水平的变化

目的：探讨叶酸缺乏对胎鼠宫内脑发育的影响，研究叶酸缺乏孕鼠子代脑组织基因组DNA甲基化水平的改变，为叶酸缺乏造成脑发育障碍提供分子和细胞水平的依据。方法：雄性SD大鼠实验组30只、对照组20只，分别饲以不含叶酸和含2mg叶酸/kg的纯合饲料，两周后与雄鼠交配，于怀孕第20天对孕鼠剖腹取胎。对模型进行评价并观察叶酸缺乏对胎鼠发育的影响，高效液相色谱法检测胎鼠脑组织基因组DNA甲基化水平。对孕鼠妊娠晚期末血清叶酸、胎鼠血清叶酸及胎鼠脑组织DNA甲基化水平作相关分析。结果：1．实验组孕鼠交配前及妊娠晚期末血清叶酸均明显低于对照组，外周血出现多分叶核粗细胞。实验组胎鼠血清叶酸也明显低于对照组，出现巨幼红细胞，RBC和Hb均低于对照组，且伴有宫内生长限制。2．实验组胎鼠脑组织基因组DNA甲基化水平低于对照组，这种改变与孕鼠及胎鼠本身的血清叶酸浓度呈正相关。结论：1．交配前两周开始限食叶酸直至妊娠期结束，所建立的叶酸缺乏孕鼠动物模型处于叶酸缺乏第三阶段，其胎鼠宫内生长受限制，红细胞出现巨幼变，但没有产生神经管闭合的异常。这是较理想的妊娠中晚期叶酸缺乏的大鼠动物模型。2．母体叶酸缺乏时胎鼠的脑组织基因组DNA甲基化水平降低，这种改变与孕鼠及胎鼠本身的血清叶酸浓度呈正相关，可能是母体叶酸缺乏时胎鼠脑发育障碍的机制之一。3．受孕前后增补叶酸宜延长至妊娠结束，以减少不良妊娠结局的危险。     

非麻醉下床边应用免缝Silo袋处理新生儿腹裂

目的 介绍非麻醉状态下于床边应用免缝Silo袋分期处理无法Ⅰ期关闭的新生儿腹裂。方法 2004年3月和7月．收治2例新生儿腹裂，分别出生后2h和4h，1例肠管污染严重，另1例合并有肠旋转不良和胎粪性肠梗阻，2例均在清醒状态下于NICU床边应用免缝Silo袋Ⅰ期处理无法回纳腹腔的肠管，2例分别于第6d和第7d在肠管完全回纳腹腔后至手术室全麻下行腹壁关闭脐部成形术，其中例2同期行Ladd’s术和肠内胎粪清除术。结果 2例患儿术后恢复好，脐部Ⅰ期愈合，分别随访5个月和2个月，生长发育好。结论非麻醉状态下床边应用免缝Silo袋分期处理无法Ⅰ期回纳的新生儿腹裂是一简便安全的方法，并可推广至产房早期处理腹裂患儿，将有效减少患儿肠管的继发性水肿和炎症，提高生存率。     

Parenteral arginine impairs intestinal adaptation following massive small bowel resection in a rat model

The nitric oxide precursor L-arginine (ARG) has been shown to influence intestinal structure and absorptive function. It is also well known that the route of administration modulates the effects of ARG. The present study evaluated the effects of parenteral ARG on structural intestinal adaptation, cell proliferation, and apoptosis in a rat model of short bowel syndrome (SBS). Male Sprague-Dawley rats were divided into three experimental groups: Sham rats underwent bowel transection and reanastomosis, SBS rats underwent a 75% small bowel resection, and SBS-ARG rats underwent a 75% small bowel resection and were treated with ARG given subcutaneously at a dose of 300 g/kg, once daily, from days 3 to 14. Parameters of intestinal adaptation, enterocyte proliferation, and enterocyte apoptosis were determined on day 15 following operation. The SBS rats demonstrated a significant increase in jejunal and ileal bowel and mucosal weight, villus height and crypt depth, and cell proliferation index compared with the sham group. The SBS-ARG animals demonstrated lower ileal bowel and mucosal weights, jejunal mucosal DNA and ileal mucosal protein, and jejunal and ileal villus height and crypt depth compared with SBS animals. The SBS-ARG rats also had a lower cell proliferation index in both jejunum and ileum and a greater enterocyte apoptotic index in ileum compared with the SBS-untreated group. In conclusion, in a rat model of SBS, parenteral arginine inhibits structural intestinal adaptation. Decreased cell proliferation and increased apoptosis are the main mechanisms responsible for decreased cell mass.     

Effect of subcutaneous insulin on intestinal adaptation in a rat model of short bowel syndrome

Insulin has been shown to influence intestinal structure and absorptive function. The purpose of the present study was to evaluate the effects of parenteral insulin on structural intestinal adaptation, cell proliferation, and apoptosis in a rat model of short bowel syndrome (SBS). Male Sprague-Dawley rats were divided into three experimental groups: sham rats underwent bowel transection and reanastomosis, SBS rats underwent a 75% small bowel resection, and SBS-INS rats underwent a 75% small bowel resection and were treated with insulin given subcutaneously at a dose of 1 U/kg, twice daily, from day 3 through day 14. Parameters of intestinal adaptation, enterocyte proliferation, and enterocyte apoptosis were determined on day 15 following operation. SBS rats demonstrated a significant increase in jejunal and ileal bowel and mucosal weight, villus height and crypt depth, and cell proliferation index compared with the sham group. SBS-INS animals demonstrated higher jejunal and ileal bowel and mucosal weights, jejunal and ileal mucosal DNA and protein, and jejunal and ileal crypt depth compared with SBS animals. SBS-INS rats also had a greater cell proliferation index in both jejunum and ileum and a trend toward a decrease in enterocyte apoptotic index in jejunum and ileum compared with the SBS untreated group. In conclusion, parenteral insulin stimulates structural intestinal adaptation in a rat model of SBS. Increased cell proliferation is the main mechanism responsible for increased cell mass.     

Lipopolysaccharide endotoxemia reduces cell proliferation and decreases enterocyte apopotosis during intestinal adaptation in a rat model of short-bowel syndrome

Sepsis is frequently associated with or complicates short-bowel syndrome (SBS). To investigate the effects of lipopolysaccharide (LPS) endotoxemia on enterocyte proliferation and death via apoptosis in a rat model of SBS, adult male Sprague-Dawley rats were divided into three experimental groups: sham rats underwent bowel transection and reanastomosis; SBS rats underwent 75% small-bowel resection; and SBS-LPS rats underwent 75% bowel resection and were given intraperitoneal injections of LPS 10 mg/kg. Parameters of intestinal adaptation (bowel and mucosal weights, mucosal DNA and protein, villus height, and crypt depth), enterocyte proliferation, and death via apoptosis were determined on day 15 after the operation. Statistical analysis was determined by Student's and ANOVA tests with a P less than 0.05 considered significant. SBS-LPS animals demonstrated a significant decrease (vs SBS rats) in duodenal (20%), jejunal (30%), and ileal (15%) overall weight, duodenal (20%), jejunal (27%), and ileal (18%) mucosal weight, jejunal (20%) and ileal (30%) mucosal DNA, jejunal (29%) and ileal (31%) villus height, and jejunal (14%) and ileal (29%) crypt depth. LPS endotoxemia led to reduced cell proliferation and enterocyte apoptosis compared to untreated SBS animals. Thus, in a rat model of SBS, LPS endotoxemia inhibits intestinal adaptation. A possible mechanism may be decreased cell proliferation. Decreased enterocyte loss via apoptosis may reflect a reduced number of enterocytes. Other mechanisms (necrosis) may be mainly responsible for cell death following LPS injection.     

Local dexamethasone improves the intestinal lesions of gastroschisis in chick embryos

Background and aims. Eviscerated bowel in gastroschisis (Gx) undergoes changes that lead to dysfunctions and create management difficulties. This study tests the hypothesis that exposure of the eviscerated bowel of chick embryos with Gx to dexamethasone might have beneficial effects on the parietal lesions.Methods. Gx was created in chick embryos on incubation day 15 and either dexamethasone (0.047 mg in 0.24 ml) or 0.075% saline were instilled into the amnio-allantoic chamber on day 17. The chicks were recovered near hatching (day 19) and eviscerated and non-eviscerated portions of the intestines were recovered, weighed and processed for HE and synaptophysin staining or for total DNA and protein measurements. Total mural and serosal layer thickness were determined and intramural ganglia were counted. ANOVA was used for comparison among groups with significance level set at p<0.05.Results. Chicks with Gx and Gx + saline controls had reduced body weight and tibial length in comparison with controls. The eviscerated bowel was heavier with marked wall thickening at the expense of all layers but particularly of the serosa. They had decreased total intestinal DNA with normal protein and decreased intramural ganglion density. In contrast, chicks from the Gx + dexamethasone group had normal body weight and tibial length, near-normal intestinal wall thickness with slightly increased serosal width, near-normal intestinal DNA content and normal density of intramural ganglia.Conclusion. Local dexamethasone had beneficial effects on the eviscerated bowel of chicks with Gx as judged by decreased wall thickening, normalization of total intestinal DNA and richer neural population. Late gestational exposure to steroids could represent another alternative for preventing intestinal lesions in Gx.     

Responsiveness of intestinal epithelial cell turnover to TGF-α after bowel resection in a rat is correlated with EGF receptor expression along the villus–crypt axis

Recent evidence suggests that transforming growth factor alpha (TGF-alpha) enhances enterocyte proliferation and stimulates intestinal adaptation after massive bowel resection. In the present study, we evaluated the effects of TGF-alpha on enterocyte turnover and correlated it with epidermal-growth factor (EGF) receptor expression along the villus-crypt axis in a rat model of short bowel syndrome (SBS). Male rats were divided into three groups, sham rats underwent bowel transection (group A); SBS rats underwent a 75% bowel resection (group B); and SBS/TGF-alpha rats underwent bowel resection and were treated with TGF-alpha (75 microg/kg) (group C) from the seventh postoperative day. Parameters of intestinal adaptation, enterocyte proliferation and apoptosis were determined on day 15. Villus tips, lateral villi and crypts were separated using laser capture microdissection. EGF receptor expression for each compartment was assessed by quantitative real-time PCR (Taqman). Statistical analysis was performed using one-way ANOVA test, with P < 0.05 considered statistically significant. Treatment with TGF-alpha resulted in a significant increase in all parameters of intestinal adaptation. EGF receptor expression in crypts significantly increased in SBS rats (vs sham rats) (0.035 +/- 0.013 vs 0.010 +/- 0.002 Log ng Total RNA/18 s) and was accompanied by a significant increase in enterocyte proliferation (169 +/- 8 vs 138 +/- 5 BrdU positive cells/per 10 crypts, P < 0.05) and decreased apoptosis following TGF-alpha administration (group C). A significant decrease in EGF receptor expression at the tip of the villus (0.005 +/- 0.002 vs 0.029 +/- 0.014 Log ng Total RNA/18 s) and in the lateral villus (0.003 +/- 0.001 vs 0.028 +/- 0.006 Log ng Total RNA/18 s) in SBS (group B) rats (vs sham, group A) was accompanied by increased cell apoptosis in these compartments following treatment with TGF-alpha (group C). In a rat model of SBS, TGF-alpha increased enterocyte proliferation and stimulated intestinal adaptation. The effect of TGF-alpha on enterocyte turnover is correlated with EGF receptor expression along the villus-crypt axis.     

Effect of leptin on intestinal re-growth following massive small bowel resection in rat

Recent evidence suggests that the adipose tissue-derived cytokine leptin (LEP) is involved in modulation of growth and differentiation of normal small intestine. The purpose of the present study was to evaluate the effects of parenteral LEP on structural intestinal adaptation, cell proliferation and apoptosis in a rat model of short bowel syndrome (SBS). Male Sprague-Dawley rats were divided into three experimental groups: Sham rats underwent bowel transection and re-anastomosis, SBS-rats underwent a 75% small bowel resection, and SBS-LEP-rats underwent bowel resection and were treated with LEP given subcutaneously at a dose of 20 μg/kg, once daily, from day 3 through 14. Parameters of intestinal adaptation (bowel and mucosal weights, mucosal DNA and protein, villus height and crypt depth in jejunum and ileum), enterocyte proliferation and enterocyte apoptosis were determined on day 15 following operation. Ileal tissue samples were taken for detection of bax and bcl-2 gene expression using RT-PCR technique. Statistical analysis was performed using the non-parametric Kruskal–Wallis ANOVA test, with P<0.05 considered statistically significant. Treatment with subcutaneous LEP resulted in a significant increase in jejunal (17%, P<0.05) and ileal (13%, P<0.05) bowel weight, jejunal (10%, P<0.05) and ileal (25%, P<0.05) mucosal weight, jejunal (26%, P<0.05) and ileal (38%, P<0.05) mucosal DNA, ileal (25%, P<0.05) mucosal protein, jejunal (41%, P<0.05) and ileal (21%, P<0.05) villus height, jejunal (37%, P<0.05) crypt depth, and jejunal (24%, P<0.05) and ileal (21%, P<0.05) enterocyte proliferation compared to SBS-animals. Enterocyte apoptosis increased significantly after bowel resection in jejunum and ileum compared to sham animals and was accompanied by an increased bax gene expression and a decreased bcl-2 gene expression in ileal samples. SBS-LEP rats showed a trend toward a decrease in enterocyte apoptosis in ileum and a mild decrease in bax gene expression compared to SBS-untreated animals. In conclusion, in a rat model of SBS parenteral LEP stimulates structural intestinal adaptation. Increased cell proliferation and decreased cell death via apoptosis may be responsible for this increased cell mass.     

Transperitoneal exchanges of water and solutes in the fetus with gastroschisis. Experimental study in the chick embryo.

Aiming at testing the hypothesis that, in the foetus with gastroschisis (GX) peritoneal dialysis during antenatal bowel exposure to the amniotic fluid may induce some modifications of the internal environment, we have studied the composition of blood, amniotic fluid and allantoic fluid in a chick embryo model of GX. Intestinal loops were surgically eviscerated in 184 embryos on the 14th day of incubation and exposed to the mixture of both embryonal fluids until the 19th day (hatching expected at the 21th). Typical lesions of GX were present in half of the 94 survivors (n = 42). For comparison we used a control group of non-manipulated animals (n = 19) and a "sham" group of animals in which the operation was carried out only until umbilical manipulation but without bowel exposure (n = 22). Chicks with GX were smaller and slightly dehydrated as attested by clinical signs and had moderately increased hematocrit. They had significant hyperkaliaemia and slight decreases in serum Na and Cl. Urea, total protein and protein fractions were not modified. Taking into account the gradients for such substances existing between the dialysate (amino-allantoic mixture) and the embryonal blood, the changes observed, albeit slight, suggest that there was some evidence of dialysis through the peritoneum: lower Na and Cl concentrations in the dialysate than in blood, induced slight falls in serum Na and Cl whereas higher K concentration resulted in severe hyperkaliemia. The high levels of albumin in the chick amniotic fluid did not allow protein depletion and therefore total protein and its fractions remained unchanged.(ABSTRACT TRUNCATED AT 250 WORDS)