2002年 · 第115卷第05期

Objective To investigate the proliferation and differentiation of cultured corneal stem cells and determine the effect of corneal stem cells cultured on amniotic membranes on the limbal area for treating corneal burns. Methods The proliferation and differentiation of corneal stem cells in vitro had been examined using colony-forming efficiency and immunohistochemistry. The stem cells had been cultured on amniotic membranes and transplanted to the limbal area for treating corneal burns. Results Corneal stem cells had a high proliferation capacity in primary and first passage, cytokeratin 3 was not expressed in primary culture but partly in first passage. The stem cells could proliferate to form cell layer on an amniotic membrane. When transplanted, stem cells could survive on limbus. After transplantation, ocular inflammation resolved, the cornea re-epithelialized, the stromal opacity reduced, the superficial neovascularity was lessened and the conjunctival fornix re-established. Conclusions Ocular surface conditions could be improved by allograft of corneal stem cells cultured on amniotic membranes. Chin Med J 2002; 115(5): 767-769

Objective To investigate whether transdermal scopolamine increased cardiac vagal activity in patients during the acute phase of myocardial infarction. Methods 30 patients with a first acute myocardial infarction and preserved sinus rhythm who were on no drug that could influence the sinus node were randomly assigned to either treatment group or placebo group. Measures of heart rate variability (HRV) in patients given drug or placebo were obtained by digital 24 hour Holter recording before and after treatment. Baroreflex sensitivity was performed using the phenylephrine method. Results No significant differences was found in the indices of the time domain and the frequency domain in both groups before treatment. Patients with transdermal scopolamine showed a significant increase in the standard deviation of normal RR intervals (SDNN), standard deviation of all five min mean normal RR intervals (SDANN), root mean square of differences of successive normal RR intervals (rMSSD), total power (TP, 0.00-0.40 Hz), low frequency peak (LF, 0.04-0.15 Hz), high frequency peak (HF, 0.15 -0.40 Hz), and Baroreflex sensitivity after treatment (P <0.05 - 0.01) . These indices did not change in patients given placebo. Conclusion Low doses of transdermal scopolamine safely increase cardiac parasympathetic activity and improve autonomic indices in patients with acute myocardial infarction. Chin Med J 2002; 115(5): 770-772

Objective To understand the distribution of prevalence rate of osteoporosis in the middle-aged and elderly in parts of China. Methods Bone mineral density (BMD) was measured and questionnaires were taken for 5593 people aged above 40 years in five administrative areas in China selected by the stratified-multi-steps-cluster sampling method. Results The total prevalence rate of osteoporosis was 16.1%. The prevalence rate among males was 11.5%and among females was 19.9%(P<0.01). There were also osteoporosis prevalence differences among cities, age groups, gender groups and areas. Conclusion The prevention and treatment of osteoporosis are very important for females but also should not be ignored in males. Chin Med J 2002; 115(5): 773-775

Objective To investigate the effect of rehabilitation therapy for short bowel syndrome on patient nutritional status and intestinal adaptation. Methods The rehabilitation therapy included enteral or parenteral nutrition, glutamine, recombinant human growth hormone and rehabilitative diet. From January 1997 to July 2000, twenty-seven patients with short bowel syndrome received the treatment. The average age of the patients was 38.5 ± 19.3 years, and the length of residual small intestine ranged from 15 to 80 cm, with an average of 46.8 ± 23.4 cm. The ileocecal valve was preserved in 14 cases, and the average time between the onset of short bowel syndrome and the rehabilitation therapy was 86 ± 105 days. Results After the treatment, nutritional status of the patients improved markedly, and intestinal absorptive capacity improved. Eight patients were followed up for more than 2 years, among whom 4 (50%) were weaned from total parenteral nutrition . Thirteen patients were followed up for more than 1 year, and 10 patients (76.9%) were weaned from total parenteral nutrition. Conclusions Rehabilitation therapy for short bowel syndrome can improve patient nutritional status effectively and promote intestinal adaptation, providing a new hope for these patients. The therapeutic effects are related to the length of the residual small intestine, patient's age and duration between massive intestinal resection and start of the treatment. Early initiation of rehabilitation therapy promotes intestinal adaptation and increases patient's ability to wean from total parenteral nutrition. Chin Med J 2002; 115(5): 776-778

Objective To investigate whether direct administration of adenoviral vectors (Ad) containing the complementary deoxyribonucleic acid (cDNA) of vascular endothelial growth factor 165 (Ad-VEGF165)induces porcine coronary collateral vessel formation, improves regional myocardial perfusion and function and is safe. Methods Three weeks after miniature swine underwent left thoracotomy and placement of an Ameroid constrictor on the left circumflex coronary artery (LCX), Ad-VEGF165 (n=6) or the control, Ad expressing β-galactosidase cDNA (Ad-Gal, n=6), was directly administered into the ischemic myocardium in the circumflex distribution. All animals were sacrificed 4 wk after the second surgery. Myocardial perfusion and function were assessed by electrocardiogram-gated single photon emission computed tomography(GSPECT) imaging. Ex vivo coronary angiography was performed to examine collateral vessels. Toxicity was assessed by blood analyses on the day just before (day 0) and on day 1, 3, 7, 28 after vector delivery and by vascular, myocardial and liver histology after sacrifice. Results GSPECT imaging 4 wk after administration of Ad-VEGF165 demonstrated significant reduction in ischemic area (P <0.01) and rest ischemic severity (P <0.01) and significant improvement in the left ventricular ejection fraction (P<0.01) and regional wall motion (P<0.05) compared with that of Ad-Gal and before administration of Ad-VEGF165. Collateral vessel development assessed by coronary angiography was significantly greater in the Ad-VEGF165 group than in the Ad-Gal group (P <0.05). General safety parameters, including routine blood parameters, liver and kidney function and cardiac specific parameters demonstrated no difference between Ad-VEGF165 and Ad-Gal animals except for the red blood cell count on day 28 (P <0.05) and blood urea nitrogen on day 7 (P <0.05). Only transient elevations in creatine phosphokinase (P <0.05) and aspartate transaminase (P <0.05) on day 1 were revealed compared with that before vector administration in both groups. Histologically, no atherosclerotic lesion in the circumflex and no inflammation in liver were revealed and only a small myocardial necrosis was observed in one Ad-VEGF165 animal (area≤20%) and one Ad-Gal animal (area <10%). Conclusions Ad-VEGF165 can induce coronary collateral vessel formation, improve regional myocardial perfusion and function and is safe by means of direct injection, which suggesting that this strategy may be useful in treating human ischemic heart disease. Chin Med J 2002; 115(5): 643-648

Objective To determine the biotic effects of angiotensin Ⅱ (Ang Ⅱ) on the migration of rat smooth muscle cells (VSMCs) and investigate the mechanisms involved in the development of vascular injury. Methods VSMCs isolated from aortic media of Wistar rats and cultured by the modified explant method were adopted. In the presence and absence of Ang Ⅱ, the expression of Ang Ⅱ receptor (ATR) and reorganization of the actin cytoskeleton and focal adhesion of VSMCs were studied by an immunocytochemistry technique and fluorocytochemistry technique. Migration assays were performed with a modified Boyden's chamber. The effects of AT1 R antagonist (CV-11974), AT2R antagonist (PD123319)on the aforementioned target were studied. Results VSMCs migration was stimulated by adding Ang Ⅱ. The dynamic reorganization of actin cytoskeleton and focal adhesions may be an important mechanism by which Ang Ⅱ facilitates VSMCs motility. The expression of AT1 R in VSMCs could be upregulated initially after treatment with Ang Ⅱ, then decreased gradually. The expression of AT1R was downregulated by AT1,R antagonists. The effect of AngⅡ on VSMCs migration was mediated by AT1R, while AT2R had no significant effect. Conclusions The dynamic reorganization of focal adhesions and the actin cytoskeleton is required for AngⅡ - induced VSMCs migration. This effect is mediated by AT1R. Chin Med J 2002; 115(5): 649-653

Objective To verify the linkage of the candidate regions identified in a previous study (markers D2S168,D2S151, D2S142 on chromosome 2) with hypertension in Chinese families. Methods A genetic linkage study focused on chromosome 2 was performed on 240 Chinese families containing 856 patients with essential hypertension. A total of 1080 individuals were genotyped using 9 highly polymorphic microsatellite markers around the candidate regions on chromosome 2 with an average spacing of 5 cM. Non-parametric linkage (NPL), parametric linkage analysis and transmission-disequilibrium test (TDT) with the GENEHUNTER software were used to assess evidence for linkage. Results Linkage of a region on chromosome 2 around D2S151 and D2S142 with hypertension was confirmed by different statistical methods (NPL, LOD score and TDT). However, the linkage of D2S168 could not be replicated in this extension study. Conclusions The data suggest that a region on chromosome 2 at or near the loci of D2S142 and D2S151 may harbor genes responsible for the development of essential hypertension in Chinese. Chin Med J 2002; 115(5): 654-657

Objective To observe the characteristics of coronary microvascular lesions (CML) in the autopsied elderly cases with hypertensive left ventricular hypertrophy (LVH) and the difference of CML among the groups of essential hypertension (EHT), coronary heart disease(CHD) and diabetes(NIDDM) also with LVH. Methods A retrospective study was performed in 206 cases≥60 years old of EHT, CHD and NIDDM with LVH and 30 normal cases as control, out of 3195 consecutive autopsied cases from 1954 to 1996 in our hospital. Arterioles with diameters of 10 - 60 μm and capillaries in the muscular layer were shown by the methods of HE, Elastic fiber + VG staining and immunohistochemistry of CD31. Quantitative measurements on the arteriole density (AD), the ratio of arteriolar wall and cavity (RWC), capillary density (CD) and the area of endothelial cell (AEC) were performed with light microscope observation and image analysis by computer. According to the thickness of the left ventricle free wall, the severity of LVH was divided into four degrees from 0 to Ⅲ . LVH of degree 0- Ⅲ was observed in EHT group, while only LVH of degree I was found in CHD, EHT + CHD, and NIDDM groups. SAS system was used for statistical analysis. Results AD and RWC increased while CD and AEC decreased significantly with the progression of LVH in EHT groups (P <0.05- 0.01). There was a similar but more severe change in the (HT + CHD) group (P <0.01); the AD increased (P <0.05) while all other measurements did not show obvious changes in the CHD group. The AD increased, CD and AEC decreased (all P <0.05), but RWC did not change very much in the NIDDM group. Conclusion CML in the EHT group was characterized by an increased AD and RWC, decreased CD and AEC, among which the increased RWC was the typical change in EHT groups compared with the groups of CHD and NIDDM. Damaged CML may be one of the main factors for decreased coronary flow reserve and myocardial ischemia in cases of EHT with LVH. Chin Med J 2002; 115(5): 658-663

Objective To investigate changes in the expression of sarcoplamic reticular Ca2+- ATPase (SERCA) and IP3-I receptors (IP3R1,) mRNA in patients with atrial fibrillation. Methods Thirty-eight patients with mitral stenosis undergoing open heart surgery were studied. 100 mg of atrial tissue was obtained during surgery from the right appendage and the right atrium. The amount of messenger ribonucleic acid (mRNA) amount of SERCA and IP3R1, was measured by reverse transcription-polymerase chain reaction (RT-PCR) and normalized to the mRNA levels of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) . Results Levels of mRNA expression of SERCA in patients with AF, as compared with subjects in sinus rhythm, was lower and that of IP3R1, was higher. The longer AF was sustained, the higher the levels of mRNA. There was no significant difference between right atrial free wall and right appendage. Conclusions The expression changes of SERCA and IP3R mRNA may correlate with the initiation or maintenance of AF. Chin Med J 2002; 115(5): 664-667

Objective To investigate the molecular mechanism of human ether-a-go-go-related gene (HERG)potassium channels regulated by protein kinase A (PKA) in a human cell line. Methods HERG channels were stably expressed in human embryonic kidney (HEK) 293 cells, and currents were measured with the patch clamp technique. The direct phosphorylation of HERG channel proteins expressed heterologously in Xenopus laevis oocytes was examined by 32P labeling and immunoprecipitation with an anti-HERG antibody. Results Elevation of the intracellular cAMP-concentration by incubation with the adenylate cyclase activator, forskolin (10 μmol/L), and the broad range phosphodiesterase inhibitor, IBMX (100 μmol/L),caused a HERG tail current reduction of 83.2%. In addition, direct application of the membrane permeable cAMP analog, 8-Br-cAMP (500 μmol/L), reduced the tail current amplitude by 29.3%. Intracellular application of the catalytic subunit of protein kinase A (200 U/ml) led to a tail current decrease by 56.9%and shifted the activation curve by 15.4 mV towards more positive potentials. HERG WT proteins showed two phosphorylated bands, an upper band with a molecular mass of ≈ 155 kDa and a lower band with a molecular mass of ≈135 kDa, indicating that both the core- and the fully glycosylated forms of the protein were phosphorylated. Conclusions PKA-mediated phosphorylation of HERG channels causes current reduction in a human cell line. The coupling between the repolarizing cardiac HERG potassium current and the protein kinase A system could contribute to arrhythmogenesis under pathophysiological conditions. Chin Med J 2002; 115(5): 668-676

Objective To investigate the course of coronary artery lesions caused by Kawasaki disease, and the value of coronary angiography (CAG) and two-dimensional echocardiography (2-D Echo) in the evaluation and follow-up of coronary artery lesions. Methods Eighty seven patients with coronary artery lesions caused by Kawasaki disease from 1979 to 1997 were retrospectively analyzed. One hundred and sixty-seven CAGs were performed in 87 patients during follow-up. CAG was repeated every 1 - 3 years in each patient until complete regression was confirmed. 2-D Echo was performed before CAG each time. The longest period of follow-up was 16 years and 6 months. Patients were treated with aspirin or aspirin and warfarin. Results During follow-up, the coronary artery lesions regressed in 48/87 (55%) patients, however, they developed into severe coronary artery lesions in 6/87 (7%) patients in whom coronary artery bypass surgery was performed. The coronary artery aneurysm regressed in some patients, while stenotic lesions remained or developed. The ratio of coronary artery stenotic lesions to aneurysms increased progressively. This study showed that Echo diagnosis of coronary artery lesions has " false positives" and false negatives" . Only 76%of coronary aneurysms and 18%of stenotic lesions could be found by 2-D Echo. No stenotic lesion could be found in distal segments of the coronary artery. Conclusions Long term follow up revealed spontaneous regression occurred in 55%of patients and development into severe coronary artery stenosis in 7%. It is necessary to perform long-term follow-up in patients with coronary artery lesions caused by Kawasaki disease. 2-D Echo can not completely replace CAG during follow-up of coronary artery lesions caused by Kawasaki disease. Chin Med J 2002; 115(5): 681-684

Objective To explore the role of nuclear factor-KB (NF-κB) in the signal conduction of protein kinase C(PKC) regulated proliferation, apoptosis and expression of Th2 cytokines-interleukin-4 (IL-4) and interleukin-5 (IL-5) of T lymphocytes in the bronchial alveolus lavage fluid (BALF). Methods T lymphocytes were isolated and purified from BALF of asthmatic guinea pigs in normal and asthmatic groups, and were stimulated with PKC agitator phorbol 12-myristate 13-acetate (PMA) and NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC), respectively. The expressions of NF-κB, IL-4 and IL-5 mRNA and protein, the proliferation and apoptosis of T lymphocytes were observed by immunohistochemistry, in situ hybridization, ELISA, MTT and TUNEL, respectively. Results The activation of NF-kB, proliferation response, and expression of IL-4 and IL-5 mRNA and protein in T lymphocytes stimulated by PMA were significantly higher than those of their blank control (P < 0.01), while those indexes of T lymphocytes stimulated by PMA and PDTC simultaneously were significantly lower than those stimulated by PMA alone (P <0.01). The apoptotic index of T lymphocytes stimulated with PMA were significantly lower than that of their blank control (P <0.01), and the apoptotic index of asthmatic guinea pig T lymphocytes stimulated with PMA and PDTC simultaneously were significantly higher than that stimulated by PMA alone (P <0.01). The significant positive correlations were found between the activation of NF-κB and the proliferation (r=0.64, P <0.001), and the expression of IL-4 and IL-5 mRNA and protein of T lymphocytes, respectively (r=0.55-0.68. P <0.001) . There was also significant negative correlation between the activation of NF-κB and apoptosis of T lymphocytes (r= 0.62, P<0.001). Conclusions NF-κB may participate in the signal conduction of PKC regulated proliferation, apoptosis and expression of IL-4 and IL-5 of T lymphocytes in asthma. The activation of NF-κB in PKC signal conduction pathway of T lymphocytes may play an important role in the pathogenesis of asthma. Chin Med J 2002; 115(5): 685-689

Objective To study the relationship between neuropeptide Y (NPY) and diabetes by examining the content and distribution of NPY and its mRNA expression in the hypothalamus and pancreas of STZ-diabetic rats. Methods Thirty Wistar rats were randomly divided into 3 groups (diabetic group, diabetic insulin treatment group, and control group). After feeding for 24 weeks, the rats were sacrificed. The expression of NPY in the hypothalamus and pancreas was detected with immunohistochemisty and in situ hybridization. Results (1) The hypothalamic content of NPY and its mRNA were significantly increased in STZ-diabetic rats in comparison with normal controls. Increased expression of NPY mRNA was found only in the arcuate nucleus and not in the paraventricular nucleus in diabetic rats, suggesting that NPY was produced in the arcuate nucleus. (2) The hypothalamic content of NPY and its mRNA in STZ-diabetic rats were visibly reduced after insulin treatment compared with that in untreated diabetic rats. This supports the hypothesis that insulin deficiency in the brain may be responsible for increased hypothalamic NPY gene expression in diabetic rats. (3) The increase of hypothalamic NPY in STZ diabetic rats associated with hyperphagia and polydipsia could be reversed by insulin replacement, suggesting that increased hypothalamic NPY contributes to the pathophysiological progress of the diabetic state. (4) The present study demonstrated for the first time that the content of NPY and its mRNA in the pancreas was increased in STZ-diabetic rats, and that the distribution of NPY-positive cell in islets was changed from the periphery to the whole islet. The content and distribution of NPY and its mRNA in islets were not changed by insulin treatment. Conclusion Increased NPY in the hypothalamus results in hypophagia and polydipsia, while the implication of increased NPY in the pancreas of diabetic rats is not clear. Chin Med J 2002; 115(5): 690-695

Objective To explore the mechanism underlying changes in microvascular reactivity in single- and double-transgenic mice. Methods Peripheral vascular reactivity to the vasodilators, acetylcholine and sodium nitroprusside, on perfused microvasculature of the hind footpad was investigated using nontransgenic mice, single-transgenic mice expressing the human APP-C100 (TgC100. WT or TgC100. V717F) and double-transgenic mice coexpressing human APP-C100 and human SOD1(G93A) genes. Results Single TgC100 and double Tg mice C100/SOD1(G93A) at 2-3 months old showed a statistical decrease of 28%in blood flux compared to nontransgenic control mice. In addition, vasodilative responsiveness was markedly reduced to 34%in 8-9 months old TgC100 mice compared to control mice. There was no significant difference in the profile of vasodilative reaction between TgC100. WT and TgC100.V717F mice. TgC100 and double Tg mice also had higher levels of Aβ peptide in plasma than nontransgenic mice (P <0.01). Conclusions The present study suggests that the altered reactivity of the microvasculature may be mediated by circulating soluble Aβ peptides. The mechanisms underlying the vasoactivity of circulating Aβ in TgC100 and double Tg mice may involve both the endothelium and nonendothelium. Chin Med J 2002; 115(5): 696-701