Compound brain peptide gangiioside can improve intrauterine hypoxia-induced neonatal brain injury and promote synapse regeneration in a mouse model / 中国组织工程研究

BACKGROUND: The most current application of compound porcine cerebroside and gangiioside injection (CPCGI) focuses on the clinical therapeutic efficacy in some craniocerebral injuries and neonatal hypoxic ischemic encephalopathy, but the molecular mechanism of CPCGI involved in the recovery of nerve function is rarely reported. OBJECTIVE: To study the protective effect and mechanism of CPCGI in a neonatal mouse model of intrauterine hypoxia-induced brain injury. METHODS: Fifteen Kunming mice were randomly divided into control group, hypoxia group and treatment group. At the 14th day after pregnancy, mice in hypoxia group and treatment group were put into a 10% oxygen incubator to make the model of intrauterine hypoxia. After delivery, neonatal mice were intraperitoneally injected with CPCGI and PBS respectively, and mouse development in each group was observed after treatment. Mice in the control group were not hypoxic and treated. The expression of glial fibrillary acidic protein, Neurocan, SynDIGI and collagen IV were observed by immunofluorescence staining and western blot assay. A step-down test was used to test animal memory function. The study protocol was approved by the Animal Ethic Committee of Luohe Medical College in China. RESULTS AND CONCLUSION: Compared with the control group, the expressions of glial fibrillary acidic protein and Neurocan in the hypoxia group were significantly increased, while the expressions of SynDIGI and collagen IV were significantly decreased. The expression of SynDIGI and collagen IV in the treatment group was significantly increased than that in the hypoxia group, while the expression of glial fibrillary acidic protein and Neuroncan was significantly decreased than that in the hypoxia group. The learning and memory ability of mice in the hypoxia group was significantly decreased compared with the control group, but was significantly improved after CPCGI treatment. These results suggest that CPCGI can alleviate brain injury and improve learning and memory ability after hypoxic ischemic encephalopathy. The detailed mechanism might be related to inhibiting activation of astrocytes, down-regulating Neuroncan expression, up-regulating collagen IV expression, promoting synaptic reconstruction and reducing cerebrovascular injury.

Therapeutic Effect of Arginyl-Fructosyl-Glucose on Diabetic Nephropathy Mice and Correlation of Gene TGF-β1 and Collagen-IV / 中国药学杂志

OBJECTIVE: To investigate the mechanism and the protective effect of arginyl-fructosyl-glucose(AFG) on the kidney of diabetic nephropathy(DN) mice and the correlation between TGF-β1 and collagen . METHODS: Twelve mice were randomly selected as control group; all others were induced by streptozotocin (STZ). The mice were randomly divided into 5 groups, each of which has 12 mice, DN model group, metformin group, 10,20,40 mg•kg-1 AFG group; each group was treated for 5 weeks, during when mice were weighed weekly and blood glucose concentration were measured. At the end of 5 weeks, the final body weight and the renal index of the mice was calculated; 24 h urinary protein and blood biochemical indexes were recorded (TC, TG, HDL-C, SCr, BUN, SOD, GSH, CAT, T-AOC); the pathological changes of the kidney was observed using HE staining. The expressions of TGF-β1 and collagen mRNA in kidneys were detected by real-time fluorescence quantitative PCR (RT-PCR). RESULTS: The levels of urinary protein, TC, TG and HDL-C in the AFG group were significantly lower than those in the control group. The contents of SOD, GSH, CAT and T-AOC in the AFG-treatment group were significantly higher than those in the control group. Pathological condition was relieved; the mRNA expression of TGF-β1 and collagen in the kidneys of each dose group was significantly lower than that of the control group. CONCLUSION: AFG protects the kidneys by reducing the total amount of 24 h urinary protein, decreasing renal pathological damage and decreasing the expression of TGF-β1 and collagen genes in kidney.

A study on prothrombogenic and antithrombogenic biomarkers in deep vein thrombosis following meta-epiphyseal cancellous bone traumatization in major orthopedic surgeries.

Background: This study was aimed to show differences in the mean values of Collagen I (Col I)/Nitric Oxide (NO), Collagen IV (Col IV)/NO, Tissue Factor (TF)/NO, and P-selectin (P-sel)/NO ratios between patients with DVT and those without DVT, following hip and knee meta-epiphyseal cancellous bone traumatization in major orthopedic surgeries. Methods: This is an observational prospective cohort study on 69 patients aged > 50 years, who had orthopedic surgery without thromboprophylaxis. Examination of serum Col I, Col IV, TF, P-sel, and NO biomarker levels were performed three times, i.e. before surgery, 72 hours and 144 hours after surgery. We looked for the differences in mean levels of biomarkers, and mean ratio values of the prothrombogenic/antithrombogenic (Col I/NO, Col IV/NO, TF/NO, P-sel/NO) at 72 hours and 144 hours post surgery between patients with DVT and those without. DVT events, which were confirmed at 144 hours post surgery by venography (with the exception of 8 cases where color Doppler ultrasound was done due to contrast usage contraindications). Results: DVTs were identified in 18 patients (26.1%). There were significant differences of mean levels in pre-surgical Col IV (p = 0.022) and 72 hours NO (p = 0.014) between patients with and without DVT. In addition, between the same two patient groups, significant differences were found in the mean values of the prothrombogenic/antithrombogenic ratios, i.e. Col IV/NO, TF/NO, and P-sel/NO at 72 hours post-surgery (p = 0.007, p = 0.028, and p = 0.049 respectively), with lower median values that were found in subjects with DVT. At 144-hours post surgery, the only significant ratio difference between the two groups was the mean values of Col IV/NO ratio (p = 0.014) with the median values that were higher than the median values at 72-hours post surgery . Conclusion: The incidence of DVT following traumatization of the meta-epiphyseal cancellous bone after major orthopedic surgeries in hip and knee is influenced by the balance of prothrombogenic and antithrombogenic factors as shown by the significant differences in Col IV/NO, TF/NO and P-sel/NO ratios at 72-hours and Col IV/NO ratio at 144 hours after surgery between DVT positive and DVT negative patients.

Effect of blood glucose fluctuation and the sustained high blood glucose on renal pathological change and collagen IV expression in diabetic rats / 中南大学学报(医学版)

Objective:To observe the effect of blood glucose lfuctuation and the sustained high blood glucose on renal pathological change and collagen IV (Col IV) expression in diabetic rats. Methods:hTe 60 male Sprague-Dawley (SD) rats were randomly assigned into a normal control group (NC) and a model group (DM). hTe rats in the normal control group were fed with normal diet, while the rats in the model group were fed with high-sucrose-high-fat diet for 6 weeks. Atfer that,streptozocin (STZ, 30 mg/kg) was injected to induce diabetic model. The model group was then randomly divided into 2 subgroups:a sustained high blood glucose group and a fluctuation blood glucose group (animals in the latter group were subcutaneously injected with insulin twice daily). Rats were sacriifced atfer 3 months and kidney tissues were dissected for HE and PAS staining, Col IV immunohistochemistry and Western blot. Results:Compared with the normal control group, the renal glomeruli and capillary basal membrane in the diabetic rats was getting larger and thicker, respectively;the capsular space and ground substance was extended and increased, respectively;the volume of renal tubule, kidney hypertrophy index, glomeruler sclerosis index and Col IV content were all increased in the diabetic rats (P Conclusion:The capillary basal membrane of kidney in diabetic rats is thicker and the ground substance is increased. The degree of glomeruler sclerosis is more serious in the blood glucose lfuctuation group compared with the sustained high blood glucose group, which is conifrmed by the increased level of Col IV.

Expression of alpha-smooth muscle actin and collagenIV in villous stroma of placental terminal villi and congestion fetal capillary in growth restricted pregnancies with severe preeclampsia / 대한산부인과학회잡지

OBJECTIVE: Preeclampsia is the major cause of prenatal mortality and morbidity. The functional disorder of uteroplacental insufficiecy is caused by the impaired uteroplacental blood flow and diffusion barrier in the villi. Functional disorders like placental circulating disorders results in morphological changes of terminal. villi as functional unit of placenta. We studied to investigate the differences in villous stoma of placental terminal villi and fetal capillary between growth restricted pregnancies with severe preeclampsia and normal preterm pregnancies. METHOD: Terminal villi was examined using light microscopy and by immunohistochemical localization of matrix molecule (alpha-smooth muscle actin and collagenIV) and the immunoreactivity of alpha-smooth muscle actin and collagenIV were evaluated in 17 cases of severe preeclampsia with intrauterine growth restriction as a study group and in 17 cases of gestational age matched normotensive preterm pregnancies as a control group. Fetal capillary congestion in terminal villi was also evaluated by Hematoxylin-Eosin staining. The patterns of immunohistochemical staining were all determinated in a visual qualitative manner (0-25%: -, 25-50%: +, 50-75%: ++, 75-100%: +++) by one pathologist. Congestion was considered to be present in a failed where the majority (>90%) of the capillaries demonstrated densely packed erythrocyte. RESULT: Expression of alpha-smooth muscle actin in the terminal villous stroma was significantly increased in study group compared with control group (P=0.0001). Expression of collagen IV in the terminal villous stroma was significantly increased in study group compared with control group (P=0.0001). Fetal capillary congestion was also significantly increased in study group compared with control group (P=0.049). CONCLUSION: The result suggests that there be the structural or biochemical difference in the villous stroma between normotensive preterm pregnancies and severe preeclampsia with intrauterine growth restriction and that the extravascular contractile system might be in the villous stroma in the severe preeclampsia intrauterine growth restriction.

Alteration in Extracellular Matrix Components in Preeclamptic Nephropathy

The preeclamptic nephropathy is characterized by swelling of endothelial cells, interposition of mesangial cells and matrix, subendothelial deposits of incompletely defined material, and thickening of the capillary walls. To determine the distribution of extracellular matrix (ECM) components in preeclamptic nephropathy, the immunohistochemical study was performed in ten renal biopsy cases using antisera to human type I, III, IV, and VI collagens, fibronectin, and laminin. In preeclamptic nephropathy, the accumulation of type IV and VI collagens, fibronectin was observed in moderate amount in the mesangium and, to some extent, in the thickened capillary walls, particularly in the subendothelial layer. In segmentally sclerotic lesions seen in six cases, the amount of type IV collagen was partly decreased, whereas those of type VI collagen and fibronectin were slightly increased. Type I collagen was expressed to a mild degree in the expanded mesangium and segmentally sclerotic lesions. The results suggest that the expression of ECM in the mesangium is increased in preeclamptic nephropathy, and the deposition of ECM components may be involved in the development and the reparative process of the characteristic glomerular lesions. The formation of sclerotic lesions may be linked to the alternative accumulation of ECM components.

Characterization of Changes in Cells and Extracellular Matrix Components during the Regenerative Processes in the Resected Rat Thyroid / 대한해부학회지

Thyroid gland is composed of follicles surrounded by basement membrane, which is known to be related to the regenerative processes in several organs. This study was performed to observe the morphological details and changes of the basement membrane components, which are the presumtive factors in thyroid regeneration, with time sequences after partial thyroidectomy and then the relations between the components and thyroid regeneration were confirmed. Thyroid galnds of adult male rats were examined before and 1, 2, 3, 4, 5, 7 and 10 days after bilateral partial[50-60%] thyroidectomy. The basememnt membranes of follicles were prominent with high eosinophilicity at later stage when H & E stained. But in PAS and EM staining, the basement membrane of the operated groups didn`t show any specific changes comparing with control group but the dense collagenous fibers at the interfollicular space outside the membrane. Immunostaining of proliferating cell nuclear antigen[PCNA], which reflects the growth fraction, showed 2.3% positivity of total follicular cells in control group. The PCNA positive ratio in experimental groups were 10.3%, 15.9%, 19.9%, 12.2%, 10.5%, 1.9%, 2.7% on postoperative 1, 2, 3, 4, 5, 7, 10 days respectively. In unilaterally thyroidectomized rats, the resected lobe revealed 18.2% of PCNA positive ratio 3 days after operation, while that in the unresected lobe was only 5.9%. This suggested the predominence of local factors in thyroid regeneration. Meanwhile, with the double immunostaining of calcitonin and PCNA discriminate between follicular cells and C-cells among the PCNA positive cells, PCNA positive C-cells were very rare. The immunostaining of laminin, fibronectin, and collagen IV showed varying intensity with the progress of regeneration. Stainability for collagen IV was maintained in all groups with some increase in 5- and 7-day groups. Laminin was well localized to the basement membrane of the control follicle, and the staining intensity for laminin was markedly increased with additional cytoplasmic staining in follicular cells in 1- to 3-day groups and then decreased to attain the control level in 10-day group. In case of fibronectin, the control follicle showed scarce staining, but the resected group showed strong positivity for fibronectin with the most intense staining in 1- to 3-day groups. Immunostaining for fibronectin still showes in 10-day group with the gradual disappearance of immunoreactivity in the center of the lobe. This study revealed the accompaning changes of the basement membrane components during the regeneration process of partially resected thyroid glands. Among them, collagen IV seems to support the structural integrity of the basement membrane. Laminin and fibronectin were supposed to be related to the thyroid follicular regeneration.

Immunohistochemical Localization of Extracellular Matrix Components in Diabetic Nephropathy

Normal human glomerular basement membrane (GBM) and mesangial matrix (MM) contain several different basement membrane components in varying degrees. The characteristic morphological and ultrastructural changes in patients with diabetic nephropathy are the thickening of the GBM and the expansion of the MM. In order to investigate the changes of extracellular matrix components in diabetes, the immunohistochemical localization was performed in 17 cases with different degrees using antisera to human collagen types I, III, IV, VI, fibronectin, and laminin. The following results were obtained: 1. The reactivity for collagen IV was increased in expanded MM in the diffuse glomerulosclerosis (GS). With the progression to the nodule formation, collagen IV was prominently decreased in the peripheral area of the nodules. 2. Collagen VI was increased in GBM and MM in the diffuse GS, it was especially prominent in the expanded MM. With the progression to nodule formation, collagen VI was prominently increased in the periphery of the nodules. 3. Interstitial collagen I and III were not stained in many of the cases with the diffuse GS. With the progression to nodule formation, these were slightly expressed. A lamellar pattern of positive reaction was noted at the periphery of the late nodular lesions. 4. Fibronectin was increased in GBM & MM in the diffuse GS, it was especially intense in the MM. With the progression to the nodule formation, the reactivity of antibody to the fibronectin was decreased. 5. Laminin was weakly stained along the GBM & trace in the MM, but was not changed in the nodular GS. In summary, the expanded mesangial matrix in the diffuse GS showed a markedly increased staining for collagen IV, fibronectin and collagen VI. Less intense linear staining for collagen VI, fibronectin, laminin, collagen IV and collagen III was noted along the GBM. In the nodular GS, the composition of the early nodules resembled that of the diffuse GS. However, the late nodular lesion of the nodular GS revealed decreased reactivity for collagen IV and fibronectin at the periphery of the nodule, where collagen VI and interstitial collagen I and III were increased in laminated pattern.

Suppressed Secretion of The Extracellular Matrix of The Cultured Corneal Epithelial Cell following Inoculation of HSV-1

In order to assess the extracellular matrix(ECM) secretion of corneal epithelial cells inoculated with herpes simplex virus-1 in vitro the indirect immunofluorescent technique was used. In this study, ECM such as fibronectin, type IV collagen and laminin were rich in cultured rabbit corneal epithelial cells under the normal condition, while those became poor after inoculation with herpes simplex virus-1. This result suggested that herpes simplex virus-1 inhibit ECM secretion of the corneal epithelial cells. These findings mean that delayed epithelial wound healing after herpes simplex virus-1 infection is due to suppressed secretion of ECM which is the principal component of corneal epithelial wound healing.