2007年 · 第120卷第16期

Abstract：Background Most epidermal cells used in skin tissue engineering are obtained from the skins of fetuses or prepuces,which can not be widely used in culturing and transplanting autologous epidermis for patients with extensive burn wounds. To solve the problem, in this study, we cultured epidermal cells from different parts of human body in vitro, and detected their growth activity.Methods Normal epidermal cells obtained from the prepuce, scalp, and axilla of male patients, were cultured and passaged. Their growth characteristics including adherent rate and growth activity were compared. Data were analyzed by homogeneity test of variance.Results In primary culture, the growth of epidermal cells from the prepuce was significantly faster than that of the epidermal cells from the scalp and axilla. In the cells obtained from the prepuce, 80% confluence was achieved on day 12, while on day 16 and day 20 in the cells from the scalp and axilla, respectively. However, no significant difference was detected in their growth and proliferation in the second passage.Conclusions Although the growth of epidermal cells obtained from the scalp and axilla is slower than that from the prepuce in primary culture, stable cell line can be established and used in preparation of auto-epidermal grafts for patients with extensive burn wounds. Therefore, the scalp and axillary skin should be considered as important sources of epidermal cells other than the prepuce。

Abstract：Background Olfactory ensheathing cells (OECs) can promote many kinds of neuron growth and axonal extension. The aim of the study was to investigate the effects of co-culturing with OECs on neuron apoptosis in vitro.Methods Apoptosis was induced by treatment of cultured dorsal root ganglion neurons with 1 mmol/L hydrogen peroxide (H2O2). Cells were randomly arranged into the following treatment groups. In group 1, OECs at different density (104/ml to 8×105/ml) were added immediately after H2O2 treatment and cells were co-cultured for 24 hours. In group 2,OECs were added at different time points (0, 4, 8, 12 and 24 hours) after H2O2 treatment. Apoptotic cell death was determined by Hoechst 33258 staining and flow cytometry (FCM). Cell viability was determined by using methyl thiazoleterazolium (MTT) assays.Results The results showed in the Hoechest 33258 staining, FCM and MTT that OECs have both the density-dependent protection and time-dependent protection on neuron apoptosis. The apoptosis decreased and the dorsal root ganglion neuron viability increased, when the density of OECs was increased in co-culture groups. But further increasing OEC density above 2×105 /ml (i.e. 8×105 /ml) failed to exert additional protection. As the interval between adding H2O2 and adding OECs was increased, the amounts of apoptosis cells were also increased. When OECs were added 24 hours after H2O2, no significant protection was observed.Conclusion These results indicated that OECs could protect dorsal root ganglion neurons from apoptosis induced by H2O2 in a density-and time-dependent manner。

Abstract：Background So far, there is no efficient treatment for pulmonary fibrosis. The objective of this study was to determine whether intramuscular injection of the hepatocyte growth factor (HGF) plasmid DNA by in vivo electroporation could prevent bleomycin-induced pulmonary fibrosis in rats, and to investigate the possible mechanisms.Methods Twenty male Wistar rats were randomly divided into four groups: control group(group C), model group (group M), early intervention group (group Ⅰ ) and late intervention group (groupⅡ). Groups M, Ⅰ and Ⅱ were intratracheally infused with bleomycin, then injected the plasmid pcDNA3.1-hHGF to group Ⅰ on day 7, 14 and 21. Group Ⅱ received the same treatment like Group Ⅰ on day 14 and 21. All the rats were killed on day 28 after bleomycin injection. We detected Homo HGF expression in the rats with ELISA method and estimated the pathological fibrosis score of lung tissue using hematoxylin eosin (HE) and Massion staining. The mRNA expression of transforming growth factor-β1 (TGF-β1),cycloxygenase-2 (COX-2), and rat HGF in rat pulmonary parenchyma were evaluated by RT-PCR.Immunohistochemistry and Western blotting were performed to determine the protein expression of transforming TGF-β1 and COX-2 in lung parenchyma.Results The plasmid pcDNA3.1-hHGF could express hHGF in NIH3T3 cells and the hHGF protein is secreted into the culture medium. The expression of hHGF protein could be monitored in quadriceps muscle, plasma and lung in Groups Ⅰ and Ⅱ. Pulmonary fibrosis levels of Groups Ⅰ and Ⅱ were obviously lower than that of group M (P＜0.05).Expression of TGF-β1 protein and mRNA in lung tissue was markedly decreased in Groups Ⅰ and Ⅱ compared with Group M (P＜0.05). The level of expression of HGF and COX-2 mRNA was higher in Groups Ⅰ and Ⅱ than in Group M (P＜0.05).Conclusions Injection of the plasmid pcDNA3.1-hHGF into skeletal muscle with electroporation has a potential role in the treatment of bleomycin-induced lung fibrosis. Exogenous HGF may inhibit the expression of TGF-β1 and regulate the crosstalk between AECs and mesenchymal fibroblasts。

Abstract：Background Sj(o)gren syndrome (SS) is an autoimmune disorder characterized by chronic lymphocytic infiltration and decreased secretion in salivary glands. Apoptosis is one of the possible mechanisms involved in acinar epithelial destruction in SS. The role of apoptosis in the initiation and effect phase of sialoadenitis is still controversial. The aim of this study was to observe the roles of apoptosis-associated proteins and serum IgG levels in sialoadenitis progression in nonobese diabetic (NOD) mice.Methods 2-, 5-, 10-, 15-, 20-week female NOD and matched BALB/c control mice were selected. Saliva and tear flow rate were measured. Serum IgG level was tested by enzyme-linked immunosorbent assay (ELISA). Number of lymphocyte foci (NLF) in submandibular glands (SMGs) was counted under routine hematoxylin/eosin-stained sections.Expression of Fas, Bcl-2 and procaspase3 proteins as well as apoptotic cells in the SMGs were detected by immunohistochemical staining and by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay respectively.Results Decreased stimulated total flow rate (STFR) and lymphocyte foci in SMGs were first observed in the 10-week NOD group. STFR was negatively correlated with NLF (P＜0.05). Serum IgG in NOD mice was significantly higher than that of the control group (P＜0.05) and showed a positive correlation with NLF (P＜0.05). Fas expression in SMGs acinar cells in NOD mice increased with age and was significantly higher compared with that in the control group. Bcl-2 expression and procaspase3 expression in SMG acinar cells in each NOD group were lower compared with those of the age-matched control mice.Conclusion Abnormal expression of Fas and Bcl-2 in the SMGs and higher level of serum IgG may contribute to the initiation of sialoadenitis and cause the glandular destruction in NOD mice。

Abstract：Background Treatment of ischemic heart disease remains an important challenge, though there have been enormous progresses in cardiovascular therapeutics. This study was conducted to evaluate whether Tongxinluo (TXL) treatment around the transplantation of mesenchymal stem cells (MSCs) can improve survival and subsequent activities of implanted cells in swine hearts with acute myocardial infarction (AMI) and reperfusion.Methods Twenty-eight Chinese mini-pigs were divided into four groups including a control group (n=7); group 2,administration of low-dose TXL alone from the 3rd day prior to AMI to the 4th day post transplantation (n=7); group 3,MSCs alone (n=7) and group 4, TXL + MSCs (n=7). AMI models were made by occlusion of the left anterior descending coronary artery for 90 minutes. Autologous bone marrow-MSCs (3×107 cells/animal) were then injected into the post-infarct myocardium immediately after AMI and reperfusion. The survival and differentiation of implanted cells in vivo were detected by immunofluorescent analysis. The data of cardiac function were obtained at baseline (1 week after transplantation) and endpoint (6 weeks after transplantation) by single photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI). Apoptosis was detected by TUNEL assay and the oxidative stress level was investigated in the post-infarct myocardium at endpoint.Results At endpoint, there was less fibrosis and inflammatory cell infiltration with more surviving myocardium in group 4 than in the control group. In group 4 the survival and differentiation of implanted MSCs were significantly improved more than that seen in group 3 alone (P＜0.0001); the capillary density was also significantly greater than in the control group,group 2 or 3 both in the infarcted zone (P＜0.0001) and the peri-infarct zone (P＜0.0001). MRI showed that parameters at baseline were not significantly different between the 4 groups. At endpoint, regional wall thickening and the left ventricular ejection fraction were increased while the left ventricular mass index, dyskinetic segments and infarcted size were decreased only in group 4 compared with control group (P＜0.0001). SPECT showed that the area of perfusion defect was significantly decreased at endpoint only in group 4 compared with control group (P＜0.0001). TUNEL assay indicated that TXL administration significantly decreased cell apoptosis in peri-infarct myocardium in groups 2 and 4. Furthermore,superoxide dismutase (SOD) significantly increased and malondialdehyde (MDA) decreased in groups 2 and 4 by the administration of TXL.Conclusions Our study demonstrates the following: (1) immediate intramyocardial injection of MSCs after AMI and reperfusion resulted in limited survival and differentiation potential of implanted cells in vivo, thus being incapable of beneficially affecting post-hearts; (2) TXL-facilitation resulted in a significant survival and differentiation potential of implanted cells in vivo via inhibition of apoptosis and oxidative stress, accompanied by significant benefits in cardiac function。

Abstract：Background So far, most of the surgical techniques for congenital blepharophimosis syndrome are two-stage procedures. In this study, we investigated a modified one-stage procedure to reduce the suffering of patients.Methods From 2003 to 2005, we adopted an one-stage technique combining blepharoptosis correction with medial canthoplasty in 16 patients with congenital blepharophimosis syndrome (10 male, 6 female; aged from 6 to 21). All the patients had bilateral severe blepharoptosis, epicanthus inversus, and flat dorsum nasi. The movement of the upper lid was 0 to 3 mm, vertical length of the eye fissure 2 to 4 mm, horizontal length 13 to 22 mm, and the distance between the eyes was 35 to 39 mm. The patients were followed up for one half to 2 years after the operation.Results In all the patients, after the operation, the horizontal length of the eyelid ＞25 mm, the vertical length ＞ 6 mm.and the distance between the eyes ＜ 35mm. The appearance of their double eyelids was satisfying.Conclusion The modified one-stage technique combining blepharoptosis correction with medial canthoplasty can achieve favorable outcomes for patients with congenital blepharophimosis syndrome。

Abstract：Background Congenital vascular rings may often cause unexplained respiratory symptoms in infants and young children. Their diagnosis and treatment are often delayed. Few studies of vascular rings have been reported in China.The aim of this study was to describe the clinical presentation, diagnosis and surgical management of infants and children with congenital vascular rings.Methods Clinical histories, physical examinations, investigations, image studies and surgical interventions were retrospectively evaluated in 7 children (age range: 2 months- 4 years, mean 7 months) with congenital vascular rings.Chest radiography was performed in all patients. Echocardiography and computed tomography (CT) with 3-dimensional (3D) reconstructions were performed in 6 patients. Esophagography, cardiac catheterization and angiography, and bronchoscopy were performed in 1, 1 and 4 children, respectively.Results Six of the 7 patients had respiratory symptoms, including recurrent cough, stridor and wheeze. Age at onset of symptoms ranged from 1 month to 11 months. Chest X-ray showed nothing important on the vascular rings, besides bronchitis and pneumonia. Contrast-enhanced CT diagnosed vascular rings in 6 patients. Four patients had double aortic arches, two had balanced arches and two were right arch dominant. One patient had a right aortic arch with left ligament and 1 patient had a pulmonary artery sling. Echocardiography failed to diagnose vascular rings in 2 patients. The esophagogram of 1 patient showed esophageal compression. Bronchoscopy of 4 patients showed compression of the distal trachea. Five of the 7 patients underwent surgical division of the vascular rings. Surgical observation confirmed the CT findings in each patient.Conclusions Patients, especially infants or young children, with recurrent respiratory symptoms such as chronic cough, stridor and wheeze, should be examined for the possible presence of congenital vascular rings.Contrast-enhanced CT can clearly show the anatomy of vascular rings. As a noninvasive technique, echocardiography is helpful for diagnosis. Early surgical management in symptomatic patients is effective。

Abstract：Background Choledochal cyst is rare in western countries. The relatively high incidence of coexistent hepatobiliary disease increases the difficulty of the surgical management of choledochal cyst. Here we analyze the diagnosis and treatment of congenital bile duct cyst in 122 Chinese adults.Methods The clinical data of 122 patients with congenital choledochal cysts admitted from 1981 to 2006 were analyzed.Results Clinical symptoms in most cases were nonspecific, resulting in delayed diagnosis. Sixty-one patients (50%) had coexistent pancreatobiliary disease. Among the 122 patients, 119 patients underwent ultrasonic examination;ERCP/MRCP was performed in 63 cases and CT in 102 cases. Abnormal pancreatobiliary duct junction was found in 48 patients. Sixteen patients had malignant lesions in the bile duct, arising in 11 of them from incomplete choledochal cyst that underwent various operations including cystenterostomy or cystojejunostomy. There was significant difference between the patients who underwent incomplete cyst resection and complete cyst resection in malignancy rate of bile duct (Chi square test, P= 0.000; odds ratio, 7.800; 95 % confidence interval, 2.450 to 24.836).Conclusions ERCP, CT and MRCP had proved their great values in the classification of the disease. Cyst excision with Roux-en-Y hepaticojejunostomy is recommended as the treatment of choice for patients with type Ⅰ or type Ⅳ cysts. For type Ⅴ cyst (Caroli's disease) with recurrent cholangitis, liver transplantation should be considered。

Abstract：Background The most frequently used therapy for post-transplantation recurrence of hepatitis B virus (HBV) infection is lamivudine, but this drug is associated with a high resistance rate due to YMDD mutant. In preliminary reports, adefovir dipivoxil (ADV) has been shown to have activity against lamivudine-resistant strains of HBV. However, clinical experience in treatment of HBV infection after liver transplantation (LT) is still not entirely clear. This study was aimed to evaluate the prophylactic efficacy of ADV plus hepatitis B immunoglobulin (HBIG) in patients with YMDD mutant before LT.Methods From March 2004 to March 2006, 16 patients with chronic hepatitis B had lamivudine-resistant YMDD mutants detected prior to liver transplantation and received treatment with ADV plus additional intramuscular HBIG after LT as prophylaxis against graft reinfection. Tests for liver function, serum HBsAg, anti-HBs (HBIG), HBeAg, anti-HBc,anti-HBe, HBV-DNA, and creatinine were assessed pre- or post-liver transplantation.Results The median follow-up of these patients post-liver transplantation was 19.4 months. Fifteen patients survived and one patient died of recurrence of hepatocellular carcinoma (HCC). There was significant difference (10.98% vs. 2.26%, P＜0.05) in YMDD mutant rate between the patients with HBV-DNA over 106 copies/ml and those with HBV-DNA less than 106 copies/ml. Fifteen patients (93.8%) had undetectable HBV-DNA at 4 weeks and 1 (6.3%) at 6 months after LT. No hepatitis B recurrence was detected by persistent testing of HBsAg, HBeAg, and HBV-DNA and no increase of serum creatinine level associated with ADV was observed in any of the patients.Conclusion ADV combined with intramuscular HBIG can effectively prevent patients with pre-transplantation YMDD mutant from HBV recurrence after LT。

Abstract：Background There is significant evidence showing that microalbuminuria and arterial compliance are sensitive markers for early cardiovascular diseases. However, whether microalbuminuria is associated with reduced arterial compliance in Chinese metabolic syndrome (MS) patients remains unknown.Methods According to the definition of MS proposed by ATPⅢ in 2001, USA, subjects (n=362) were divided into three groups according to the number of risk factors: group 1 (control), group 2 (medium, ＜ 3 risk factors) and group 3 (MS, ≥ 3 risk factors). Both large artery compliance (C1) and small artery compliance (C2) were measured with the CVProfilor DO-2020 Cardiovascular Profiling System, and microalbuminuria was evaluated with the ratio of albumin to urine creatinine.Results (1) As C1 and C2 levels elasticity decreased, albumin creatinine ratio (ACR) and the prevalence of microalbuminuria increased within those groups with MS risk factors. C1 and C2 were negatively correlated with the ranking of MS risk factors, ACR was positively correlated with the ranking of MS risk factors (all P＜0.05). (2) Subjects were also categorized into a microalbuminuria group and a normal group, C1 and C2 in the microalbuminuria group were lower than in the normal group. (3) Multivariate regression analysis showed that increased systolic blood pressure (SBP) and reduced arterial compliance were the main risk factors for microalbuminuria in the MS group.Conclusions The risk of developing microalbuminuria was higher in the subjects with multiple metabolic abnormalities.Increased systolic blood pressure and reduced arterial compliance may be the main predictors for microalbuminuria in MS。

Many kinds of cell-free dermal substitutes have been developed during the past several years, however,their biomechanical properties, including hysteresis,stress relaxation, creep, and non-linear stress-strain, are still unknown. In this study, we tested these biomechanical characteristics of four dermal substitutes,and compared them with those of fresh human skin (FHS)。

Mechanical ventilation (MV) may aggravate lung injury induced by a variety of injuries, including intratracheal hydrochloric acid instillation,1 intratracheal lipopolysaccharide (LPS) instillation with or without concurrent saline lavage,2 intravenous LPS,3 or intravenous oleic acid.4 However, the mechanism for this detrimental effect of MV is unclear。

The prognosis of pancreatic carcinoma is disappointing due to the difficulty of early and accurate diagnosis,low operative resection rate, insensitivity to radiation therapy and chemotherapy。

Neurofibromatosis type 2 (NF-2) is one of the most common single gene disorders in the nervous system.For approximately 96% of patients with NF-2 present with bilateral Schwannomas involving the eighth cranial nerves, which may be accompanied by Schwannomas involving other cranial, spinal or peripheral nerves, NF-2 is also referred to as "bilateral acoustic neuromas"。

Hypophosphatemic vitamin D-resistant rickets or X-linked hypophosphatemia (XLH) is a rare hereditary metabolic disease manifesting marked hypophosphatemia, short stature and rickets. Its prevalence is approximately 1 in 20 000. Except early exfoliation of the teeth, there are a few oral findings of XLH described in China.1,2 Here we present two cases in one family。

Situs inversus totalis, which refers to a mirror-image reversal of the normal position of the internal organs,is rare and confusing. It is often associated with other anomalies, which may disturb the surgical procedure for diseases。

Surgical resection is preferred in the treatment of gastrointestinal stromal tumor (GIST).1-3 Duodenal GIST comprises 4.5% of all GISTs,1 but the optimal surgical procedure for it remains uncertain。

Urethritis glanduris is a rare benign inflammatory lesion and has a tendency to malignant transformation. We described here a case of urethritis glandularis with the extensive lesions in the anterior urethra and the special outward appearance; no similar case has been reported previously。

The 2007 Changsha International Symposium on Lupus, co-sponsored by the Institute of Dermatovenereology and the Epigenetic Research Center of the Second Xiangya Hospital at the Central South University was successfully held in Changsha on May 27,2007。