为充分利用鱼片加工废弃物鱼骨中的胶原蛋白质,采用单因素以及正交实验法,研究胃蛋白酶在酸性介质条件下提取罗非鱼骨胶原蛋白质的最佳工艺,精制后的胶原蛋白质采用SDSPAGE电泳、紫外及傅立叶变换红外扫描,圆二色检测,氨基酸分析,热变...为充分利用鱼片加工废弃物鱼骨中的胶原蛋白质,采用单因素以及正交实验法,研究胃蛋白酶在酸性介质条件下提取罗非鱼骨胶原蛋白质的最佳工艺,精制后的胶原蛋白质采用SDSPAGE电泳、紫外及傅立叶变换红外扫描,圆二色检测,氨基酸分析,热变性温度测定等方法 ,分析胶原蛋白质的理化性质。结果显示:乳酸溶液p H 2.6,温度30℃,料液比1∶30(g/mL),加酶量200U/g,提取时间24h,此工艺条件下的粗胶原蛋白质提取率达到70.28%,得率为12.76%;罗非鱼骨胶原蛋白质含有2条α链和1条β链,属于典型的I型胶原蛋白质;罗非鱼骨胶原蛋白质的主要氨基酸为甘氨酸、脯氨酸、羟脯氨酸、谷氨酸和丙氨酸;罗非鱼骨胶原蛋白质的热变性温度为34.5℃。研究表明,采用胃蛋白酶提取鱼骨胶原蛋白质,提取率和纯度均较高。展开更多
贝壳是一种具有优异力学性能的生物硬组织,贝壳基质蛋白质对贝壳的形成具有重要意义。厚壳贻贝(Mytilus coruscus)贝壳中发现一种类似胶原蛋白质的新型贝壳基质蛋白质,命名为collagen-like protein 2(CLP-2)。然而,该蛋白质的结构与功...贝壳是一种具有优异力学性能的生物硬组织,贝壳基质蛋白质对贝壳的形成具有重要意义。厚壳贻贝(Mytilus coruscus)贝壳中发现一种类似胶原蛋白质的新型贝壳基质蛋白质,命名为collagen-like protein 2(CLP-2)。然而,该蛋白质的结构与功能以及对贝壳形成的影响机制尚不清楚。为此,本研究对CLP-2开展了序列分析;进一步采取密码子优化结合原核重组表达策略,开展了CLP-2的重组表达;在此基础上分析了重组CLP-2对酸钙结晶的诱导、结晶速率抑制以及碳酸钙结合能力。对CLP-2的序列分析结果表明,该蛋白质序列中含有信号肽及两个Von Willebrand factor A(VWA)结构域。CLP-2在数据库中尚无高同源性蛋白质存在,表明这是一种较为新颖的贝壳基质蛋白。所获得的重组CLP-2对碳酸钙体外结晶表现出明显的诱导作用,扫描电镜以及傅里叶红外光谱结果表明,重组CLP-2可诱导碳酸钙晶体的形貌由立方体形转化为球形,并在高浓度下进一步转化为哑铃形;同时,重组CLP-2可促使碳酸钙晶体的晶型由方解石型向文石型转化;重组CLP-2在体外具有碳酸钙晶体结合作用;此外,重组CLP-2能显著抑制碳酸钙晶体的结晶速度(P<0.01),并具有浓度依赖性。上述结果表明,厚壳贻贝贝壳CLP-2蛋白质在贝壳,特别是文石型肌棱柱层的生物矿化过程中具有重要作用。上述研究为深入了解贻贝贝壳的形成机制,以及胶原类蛋白质对生物矿化过程的影响奠定了基础。展开更多
A群链球菌胶原蛋白样蛋白质(Streptococcal collagen-like protein,Scl)是2000年后发现的一种A群链球菌(Group A Streptococcus,GAS)的致病因子,具有与胶原蛋白类似的特殊结构,分为Scl1和Scl2两种,也称为SclA和SclB。Scl可以与人的肺上...A群链球菌胶原蛋白样蛋白质(Streptococcal collagen-like protein,Scl)是2000年后发现的一种A群链球菌(Group A Streptococcus,GAS)的致病因子,具有与胶原蛋白类似的特殊结构,分为Scl1和Scl2两种,也称为SclA和SclB。Scl可以与人的肺上皮细胞以及人体中其他几种分子结合,如补体调节蛋白H因子、低密度脂蛋白等。本文对Scl的结构特征、功能及其编码基因作了综述。展开更多
Objective The present study aimed to explore the role of P2Y1 receptor in glial fibrillary acidic protein (GFAP) production and glial cell line-derived neurotrophic factor (GDNF) secretion of astrocytes under isch...Objective The present study aimed to explore the role of P2Y1 receptor in glial fibrillary acidic protein (GFAP) production and glial cell line-derived neurotrophic factor (GDNF) secretion of astrocytes under ischemic insult and the related signaling pathways. Methods Using transient right middle cerebral artery occlusion (tMCAO) and oxygen-glucose-serum deprivation for 2 h as the model of ischemic injury in vivo and in vitro, immunofluorescence, quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR), Western blotting, enzyme linked immunosorbent assay (ELISA) were used to investigate location of P2Y1 receptor and GDNF, the expression of GFAP and GDNF, and the changes of signaling molecules. Results Blockage of P2Y1 receptor with the selective antagonist N^6-methyl-2′-deoxyadenosine 3′,5′-bisphosphate diammonium (MRS2179) reduced GFAP production and increased GDNF production in the antagonist group as compared with simple ischemic group both in vivo and in vitro. Oxygen-glucose-serum deprivation and blockage of P2Y1 receptor caused elevation of phosphorylated Akt and cAMP response element binding protein (CREB), and reduction of phosphorylated Janus kinase2 (JAK2) and signal transducer and activator of transcription3 (STAT3, Ser727). After blockage of P2Y1 receptor and deprivation of oxygen-glucose-serum, AG490 (inhibitor of JAK2) reduced phosphorylation of STAT3 (Ser727) as well as expression of GFAP; LY294002, an inhibitor of phosphatidylinositol 3-kinase (PI3-K), decreased phosphorylation of Akt and CREB; the inhibitor of mitogen-activated protein kinase kinase 1/2 (MEK 1/2) U0126, an important molecule of Ras/extracellular signal- regulated kinase (ERK) signaling pathway, decreased the phosphorylation of JAK2, STAT3 (Ser727), Akt and CREB. Conclusion These results suggest that P2Y1 receptor plays a role in the production of GFAP and GDNF in astrocytes under transient ischemic condition and the related signaling pathways may be JAK2/STAT3 and PI3-K/Akt/CREB, respectively, and that crosstalk probably exists between them.展开更多
Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMP) play a significant role in regulating angiogenesis, the process of new blood vessel formation. Interstitial collagenase (MMP-1), 72 k...Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMP) play a significant role in regulating angiogenesis, the process of new blood vessel formation. Interstitial collagenase (MMP-1), 72 kDa gelatinase A/type IV collagenase (MMP-2), and 92 kDa gelatinase B/type IV collagenase (MMP-9) dissolve extracellular matrix (ECM) and may initiate and Promote angiogenesis. TIMP-1, TIMP-2, TIMP-3, and possibly,TIMP-4 inhibit neovascularisation. A new paradigm is emerging that matrilysin (MMP-7), MMP-9, and metalloelastase (MMP-12) may block angiogehesis by converting plasndnogen to angiostatin, which is one of the most potent angiogenesis antagonists. MMPs and TIMPs play a complex role in regulating angiogenesis. An understanding of the biochemical and cellular pathways and mechanisms of angiogenesis will provide importal information to allow the control of angiogenesis, e.g. the stimulation of angiogenesis for coronary collateral circulation formation; while the inhibition for treating arthritis and cancer.展开更多
A 76-year-old woman with a 5-mo history of recurrent diarrhea and generalized edema was admitted to our hospital. Colonoscopy revealed edematous mucosa,and histopathological examination was compatible with collagenous...A 76-year-old woman with a 5-mo history of recurrent diarrhea and generalized edema was admitted to our hospital. Colonoscopy revealed edematous mucosa,and histopathological examination was compatible with collagenous colitis. Protein leakage from the colon,particularly in the ascending portion,was identified on 99mTc-human serum albumin scintigraphy. Collagenous colitis associated with protein-losing enteropathy (PLE) without small bowel disease was diagnosed. Prednisolone treatment ameliorated diarrhea and hypoproteinemia. Collagenous colitis should be included in the differential diagnosis of chronic diarrhea with hypoproteinemia for appropriate management.展开更多
AIM:Matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) are central to the spontaneous resolution of liver fibrosis. The mechanisms involved have been investigated in hepatic...AIM:Matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) are central to the spontaneous resolution of liver fibrosis. The mechanisms involved have been investigated in hepatic stellate cells (HSC), but not in hepatocytes. We investigated the effects of fibril- and fixed-collagen on MMP-1 and TIMP-1 production in hepatocytes, using the HLE cell line. METHODS: Fibril type I and IV collagen were prepared by HCI digestion of type I and IV collagen, respectively. For fixed-collagen, culture dishes were coated with fibril type I or IV collagen and fixed by ultraviolet. Type I collagenase activity was measured using fluorescein isothiocyanate-labeled type I collagen. MMP-1 and TIMP-1 in HLE cells were measured by a one-step sandwich enzyme immunoassay. RESULTS: Both fibril type I and IV collagen significantly increased type I collagenase activity about two-fold compared with no fibril collagen. The effects of the fibril collagen were not affected by the coating condition. There was no significant difference in the effects on collagenase activity between cells cultured in medium containing fibril type I collagen and those cultured in the presence of type IV collagen. Both types of fibril collagen significantly increased MMP-1 production, and showed more than 10-fold higher levels of MMP-1 than the control. The enhanced MMP-1 production by fibril collagens was unaffected by the coating condition. By contrast, TIMP-1 production was not changed by the addition of fibril type I or IV collagen, and neither was it affected by the coating conditions. Coating with type I collagen significantly suppressed MMP-1 production by almost one-tenth compared with no coating. By contrast, TIMP-1 production was not affected by either the absence of a collagen coat or by increasing the concentration of the coating collagen. CONCLUSION: These results indicated that, in HLE cells, fibril- and fixed-collagen have opposite effects on MMP-1 production without affecting TIMP production. Fibril collagen induced collagenase activity by up-regulation of MMP-1 production without affecting TIMP-1 production. By contrast, fixed collagen reduced MMP-1 production. Our results suggest that hepatocytes might also play an important role in the regulation of the hepatic fibrosis alongside HSC.展开更多
文摘为充分利用鱼片加工废弃物鱼骨中的胶原蛋白质,采用单因素以及正交实验法,研究胃蛋白酶在酸性介质条件下提取罗非鱼骨胶原蛋白质的最佳工艺,精制后的胶原蛋白质采用SDSPAGE电泳、紫外及傅立叶变换红外扫描,圆二色检测,氨基酸分析,热变性温度测定等方法 ,分析胶原蛋白质的理化性质。结果显示:乳酸溶液p H 2.6,温度30℃,料液比1∶30(g/mL),加酶量200U/g,提取时间24h,此工艺条件下的粗胶原蛋白质提取率达到70.28%,得率为12.76%;罗非鱼骨胶原蛋白质含有2条α链和1条β链,属于典型的I型胶原蛋白质;罗非鱼骨胶原蛋白质的主要氨基酸为甘氨酸、脯氨酸、羟脯氨酸、谷氨酸和丙氨酸;罗非鱼骨胶原蛋白质的热变性温度为34.5℃。研究表明,采用胃蛋白酶提取鱼骨胶原蛋白质,提取率和纯度均较高。
文摘A群链球菌胶原蛋白样蛋白质(Streptococcal collagen-like protein,Scl)是2000年后发现的一种A群链球菌(Group A Streptococcus,GAS)的致病因子,具有与胶原蛋白类似的特殊结构,分为Scl1和Scl2两种,也称为SclA和SclB。Scl可以与人的肺上皮细胞以及人体中其他几种分子结合,如补体调节蛋白H因子、低密度脂蛋白等。本文对Scl的结构特征、功能及其编码基因作了综述。
基金the National Natural Science Foundation of China (No. 30500189)
文摘Objective The present study aimed to explore the role of P2Y1 receptor in glial fibrillary acidic protein (GFAP) production and glial cell line-derived neurotrophic factor (GDNF) secretion of astrocytes under ischemic insult and the related signaling pathways. Methods Using transient right middle cerebral artery occlusion (tMCAO) and oxygen-glucose-serum deprivation for 2 h as the model of ischemic injury in vivo and in vitro, immunofluorescence, quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR), Western blotting, enzyme linked immunosorbent assay (ELISA) were used to investigate location of P2Y1 receptor and GDNF, the expression of GFAP and GDNF, and the changes of signaling molecules. Results Blockage of P2Y1 receptor with the selective antagonist N^6-methyl-2′-deoxyadenosine 3′,5′-bisphosphate diammonium (MRS2179) reduced GFAP production and increased GDNF production in the antagonist group as compared with simple ischemic group both in vivo and in vitro. Oxygen-glucose-serum deprivation and blockage of P2Y1 receptor caused elevation of phosphorylated Akt and cAMP response element binding protein (CREB), and reduction of phosphorylated Janus kinase2 (JAK2) and signal transducer and activator of transcription3 (STAT3, Ser727). After blockage of P2Y1 receptor and deprivation of oxygen-glucose-serum, AG490 (inhibitor of JAK2) reduced phosphorylation of STAT3 (Ser727) as well as expression of GFAP; LY294002, an inhibitor of phosphatidylinositol 3-kinase (PI3-K), decreased phosphorylation of Akt and CREB; the inhibitor of mitogen-activated protein kinase kinase 1/2 (MEK 1/2) U0126, an important molecule of Ras/extracellular signal- regulated kinase (ERK) signaling pathway, decreased the phosphorylation of JAK2, STAT3 (Ser727), Akt and CREB. Conclusion These results suggest that P2Y1 receptor plays a role in the production of GFAP and GDNF in astrocytes under transient ischemic condition and the related signaling pathways may be JAK2/STAT3 and PI3-K/Akt/CREB, respectively, and that crosstalk probably exists between them.
文摘Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMP) play a significant role in regulating angiogenesis, the process of new blood vessel formation. Interstitial collagenase (MMP-1), 72 kDa gelatinase A/type IV collagenase (MMP-2), and 92 kDa gelatinase B/type IV collagenase (MMP-9) dissolve extracellular matrix (ECM) and may initiate and Promote angiogenesis. TIMP-1, TIMP-2, TIMP-3, and possibly,TIMP-4 inhibit neovascularisation. A new paradigm is emerging that matrilysin (MMP-7), MMP-9, and metalloelastase (MMP-12) may block angiogehesis by converting plasndnogen to angiostatin, which is one of the most potent angiogenesis antagonists. MMPs and TIMPs play a complex role in regulating angiogenesis. An understanding of the biochemical and cellular pathways and mechanisms of angiogenesis will provide importal information to allow the control of angiogenesis, e.g. the stimulation of angiogenesis for coronary collateral circulation formation; while the inhibition for treating arthritis and cancer.
文摘A 76-year-old woman with a 5-mo history of recurrent diarrhea and generalized edema was admitted to our hospital. Colonoscopy revealed edematous mucosa,and histopathological examination was compatible with collagenous colitis. Protein leakage from the colon,particularly in the ascending portion,was identified on 99mTc-human serum albumin scintigraphy. Collagenous colitis associated with protein-losing enteropathy (PLE) without small bowel disease was diagnosed. Prednisolone treatment ameliorated diarrhea and hypoproteinemia. Collagenous colitis should be included in the differential diagnosis of chronic diarrhea with hypoproteinemia for appropriate management.
文摘AIM:Matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) are central to the spontaneous resolution of liver fibrosis. The mechanisms involved have been investigated in hepatic stellate cells (HSC), but not in hepatocytes. We investigated the effects of fibril- and fixed-collagen on MMP-1 and TIMP-1 production in hepatocytes, using the HLE cell line. METHODS: Fibril type I and IV collagen were prepared by HCI digestion of type I and IV collagen, respectively. For fixed-collagen, culture dishes were coated with fibril type I or IV collagen and fixed by ultraviolet. Type I collagenase activity was measured using fluorescein isothiocyanate-labeled type I collagen. MMP-1 and TIMP-1 in HLE cells were measured by a one-step sandwich enzyme immunoassay. RESULTS: Both fibril type I and IV collagen significantly increased type I collagenase activity about two-fold compared with no fibril collagen. The effects of the fibril collagen were not affected by the coating condition. There was no significant difference in the effects on collagenase activity between cells cultured in medium containing fibril type I collagen and those cultured in the presence of type IV collagen. Both types of fibril collagen significantly increased MMP-1 production, and showed more than 10-fold higher levels of MMP-1 than the control. The enhanced MMP-1 production by fibril collagens was unaffected by the coating condition. By contrast, TIMP-1 production was not changed by the addition of fibril type I or IV collagen, and neither was it affected by the coating conditions. Coating with type I collagen significantly suppressed MMP-1 production by almost one-tenth compared with no coating. By contrast, TIMP-1 production was not affected by either the absence of a collagen coat or by increasing the concentration of the coating collagen. CONCLUSION: These results indicated that, in HLE cells, fibril- and fixed-collagen have opposite effects on MMP-1 production without affecting TIMP production. Fibril collagen induced collagenase activity by up-regulation of MMP-1 production without affecting TIMP-1 production. By contrast, fixed collagen reduced MMP-1 production. Our results suggest that hepatocytes might also play an important role in the regulation of the hepatic fibrosis alongside HSC.