Background:Maintaining the structural and functional integrity of membranes is essential for proper cells function.A recent proteomic study suggests that S100A16 and Annexin A4(ANXA4)proteins participate to maintain t...Background:Maintaining the structural and functional integrity of membranes is essential for proper cells function.A recent proteomic study suggests that S100A16 and Annexin A4(ANXA4)proteins participate to maintain the membrane integrity in the outer segment of the photoreceptors in the eye.The protein S100A16,recently discovered,is one of the S100 family proteins for which no protein and membrane interaction has yet been identified.Furthermore,maintain of the membrane integrity is calcium sensitive process.The main objective consists of studying the membrane interactions of S100A16 and ANXA4 proteins to better understand their functions in maintaining membrane integrity.Specific objectives are:(I)to achieve the purification of these proteins,(II)to gather information on their membrane interactions,and(III)to study the influence of calcium on these interactions.Methods:The S100A16 protein is obtained by cleavage followed by purification on a His-Trap column.Membrane interactions are studied with the Langmuir monolayer model.After measurement of the saturating concentration,the protein binding parameters,that to say the maximum insertion pressure and synergy,will then be determined in the presence of several phospholipids representative of physiological membranes.Results:The S100A16 protein was obtained with a purity greater than 99%and its saturating concentration is 0.5μM.Biophysical studies with different phospholipids in monolayer are currently in progress.Conclusions:Obtaining the S100A16 protein with a high purity allows carrying out the biophysical study in order to understand its membrane interactions.The purification of ANXA4 and the biophysical study with different phospholipids of this protein alone and in complex with the S100A16 protein will allow a better understanding of the membrane behavior of these proteins,as well as their roles in the maintenance of membrane integrity.展开更多
Background:(I)To describe the development and components of the automobile simulator driving behavior evaluation system developed by CRIR-Institut Nazareth et Louis-Braille;(II)to present the preliminary results of th...Background:(I)To describe the development and components of the automobile simulator driving behavior evaluation system developed by CRIR-Institut Nazareth et Louis-Braille;(II)to present the preliminary results of the content evaluation of the driving behavior evaluation grid.Methods:The evaluation system consists of five components:(I)the VS500M Car Simulator(Virage Simulation);(II)four VS500M driving scenarios,modified to minimize the occurrence of simulator sickness and expose subjects to commonly encountered driving situations on highways and city boulevards;(III)the Tobii Pro Glasses 2 eye tracking device;(IV)a car simulator driving behavior observation grid(DBOG);(V)a software application used during the behaviour evaluation phase,where synchronized video tracking,certain data from the simulator(e.g.,speed)and the DBOG grid are presented.Initially,the expected safe driving behaviors were identified,including 235 of a visual nature,supported by literature data and consultation of the project steering committee and an expert in driving assessment.Driving behaviors were assessed in 22 subjects without visual impairment(mean age 55±20 years).Subsequently,the items were revised to determine their relevance based on their importance in terms of road safety or on the frequency with which behaviors were observed among participants.For analysis purpose,the items of the DBOG were grouped according to their content,by type of expected driving behavior(e.g.,following a stop,look to the left and right before crossing the intersection)or element to be detected(e.g.,pedestrians).Results:Some visual behaviors are difficult to observe with the eye tracker device because they are more dependent on peripheral than central vision.Others are rarely observed,possibly because they are little or not realized in daily life or the representation of reality on the simulator does not stimulate their adoption.On the other hand,the visual detection behaviors expected in a situation where safety can be compromised are mostly carried out(e.g.,detection of oncoming vehicles,side mirror verification when changing lanes).Conclusions:This first phase of the study has made possible to better target the items to be kept in the car simulator driving behavior observation grid,and those that will have to be removed as they are too difficult to observe or too rarely adopted by the participants.Content validity and inter-rater reliability will be assessed from the simplified grid.展开更多
Background:The S100A10 protein might be an early biomarker of diabetes development leading to diabetic retinopathy.The protein complex S100A10/annexin A2 allows the recruitment of the C-terminal of AHNAK protein(AHNAK...Background:The S100A10 protein might be an early biomarker of diabetes development leading to diabetic retinopathy.The protein complex S100A10/annexin A2 allows the recruitment of the C-terminal of AHNAK protein(AHNAK C-ter peptide)to the membrane in presence of calcium,before forming a platform which can initiate membrane repair.However,no molecular data are currently available on membrane binding of the different proteins involved in this complex.We aim to study the membrane binding of S100A10,AHNAK C-ter peptide and their complex to better understand their roles in cell membrane repair process.Methods:Firstly,S100A10 will be overexpressed and purified by affinity chromatography and AHNAK C-ter peptide will be synthesized.Langmuir monolayers membrane model will then be used to characterize the interactions between these proteins and different phospholipids found in membranes.The secondary structure,orientation and membrane organization of these proteins will be studied by Polarization Modulation Infrared Reflection-Absorption Spectroscopy.Their lateral localization will be determined through the influence of these proteins on the physical state of lipids by fluorescence microscopy.Results:The optimization of the overexpression,purification and cleavage of the GST tag procedure to obtain pure S100A10 was completed.Protein identification by mass spectrometry and circular dichroism stability pre-studies were performed.In parallel,AHNAK C-ter peptide was studied by Langmuir monolayer model and the results indicate this peptide prefers lipids with negatively charged polar heads and unsaturated acyl chains.Preliminary solid-state NMR results confirm this phenomenon at 37℃.Conclusions:Our research will complete current knowledge on membrane binding of S100A10 and AHNAK C-ter peptide.We could also identify the conditions leading to modifications of these membrane bindings,and possibly to the loss of protein function.Thus,this project helps to better determine their roles in membrane repair,as well as in other physiological mechanisms in which these proteins are involved.展开更多
Background The levels of long-term elevated serum or intracellular free fatty acid （FFA） induce insulin resistance associated with central obesity. The insulin-mimetic protein visfatin is preferentially produced by ...Background The levels of long-term elevated serum or intracellular free fatty acid （FFA） induce insulin resistance associated with central obesity. The insulin-mimetic protein visfatin is preferentially produced by visceral adipose tissues and has been implicated in obesity and insulin resistance. To identify that FFA is capable of inducing insulin resistance and to clarify the role of FFA on visfatin, we examined the effect of monounsaturated FFA oleate （C 18： 1） and saturated FFA palmitate （C 16：0） on glucose transport and visfatin gene expression in cultured 3T3-L1 adipocytes or preadipocytes.Methods FFA-free DMEM/F12, 0.125 mmol/L, 0.5 mmol/1 and 1.0 mmol/L oleate or palmitate was added to cultured 3T3-L1 adipocytes or preadipocytes and incubated overnight. Glucose transport was assessed as 3H- 2-deoxy-glucose uptake. Total RNA was extracted and subjected to RT-PCR for the measurement of visfatin mRNA levels. Statistical comparisons between control group and other groups were performed with the two-tailed paired t test, and one-way ANOVA was used to compare the mean values among the groups.Results Insulin increased specific membrane glucose transport in 3T3-L1 preadipocytes. Upregulation was evident from 15 minutes to 1 hour exposure to insulin. However, after 6-hour exposure to insulin, there was a downregulation in the response to insulin. Dose response studies demonstrated that 2-deoxy glucose transport was increased by 336% at 50 nmol/L insulin （P〈0.01）, and reached a maximal effect at 100 nmol/L insulin （P〈0.01）. Oleate and palmitate treatment did not influence basal glucose transport （without insulin stimulation）, whereas insulin-stimulated glucose transport was inhibited after overnight oleate and palmitate treatment in preadipocytes and adipocytes. In 3T3-L1 preadipocytes, insulin resistance could be achieved at 0.125 mmol/L oleate or palmitate （P〈0.05, respectively）, and the inhibition was dose dependent. In adipocytes, the inhibition was noted at 0.5 mmol/L oleate or 1.0 mmol/L palmitate. Visfatin mRNA expression increased during differentiation more than 1.5-fold. Bovine serum albumin （BSA） did not influence visfatin mRNA expression compared with the control group. Dose-response studies demonstrated that addition of 0.125 mmol/L oleate and palmitate to 3T3-L1 adipocytes decreased visfatin mRNA expression significantly （78%, 77%, respectively, relative to untreated control, P〈0.05）, and further to 65% （relative to untreated control, P〈0.05） and 55% （relative to untreated control, P〈0.01） at 1.0 mmol/L FFA. Furthermore, the suppression on preadipocytes was similar to that of adipocytes, which reached a maximal reduction of 44% （oleate, P〈0.05） and 47% （palmitate, P〈0.05） at 1.0 mmol/L FFA.Conclusions Oleic acid and palmitic acid may induce insulin resistance in 3T3-L1 adipocytes and preadipocytes. Downregulation of visfatin mRNA may contribute to impair insulin sensitivity caused by oleate and palmitate.展开更多
Chronic inflammatory lung disease is a common ,health problem and often treated with potent antibiotics, anti-tuberculosis drugs, and antifungal agents. However, in case of medical therapy failure, surgical treatment ...Chronic inflammatory lung disease is a common ,health problem and often treated with potent antibiotics, anti-tuberculosis drugs, and antifungal agents. However, in case of medical therapy failure, surgical treatment has been often considered as an effective procedure. Indications for surgical procedure mainly include recurrent infection, acute suppurative complications including lung abscess and empyema, and often chronic, intermittent, or massive hemoptysis.1 Segmentectomy or lobectomy is considered as an ＂ideal＂ procedure when the disease is localized to one lung and single segment or lobe.展开更多
文摘Background:Maintaining the structural and functional integrity of membranes is essential for proper cells function.A recent proteomic study suggests that S100A16 and Annexin A4(ANXA4)proteins participate to maintain the membrane integrity in the outer segment of the photoreceptors in the eye.The protein S100A16,recently discovered,is one of the S100 family proteins for which no protein and membrane interaction has yet been identified.Furthermore,maintain of the membrane integrity is calcium sensitive process.The main objective consists of studying the membrane interactions of S100A16 and ANXA4 proteins to better understand their functions in maintaining membrane integrity.Specific objectives are:(I)to achieve the purification of these proteins,(II)to gather information on their membrane interactions,and(III)to study the influence of calcium on these interactions.Methods:The S100A16 protein is obtained by cleavage followed by purification on a His-Trap column.Membrane interactions are studied with the Langmuir monolayer model.After measurement of the saturating concentration,the protein binding parameters,that to say the maximum insertion pressure and synergy,will then be determined in the presence of several phospholipids representative of physiological membranes.Results:The S100A16 protein was obtained with a purity greater than 99%and its saturating concentration is 0.5μM.Biophysical studies with different phospholipids in monolayer are currently in progress.Conclusions:Obtaining the S100A16 protein with a high purity allows carrying out the biophysical study in order to understand its membrane interactions.The purification of ANXA4 and the biophysical study with different phospholipids of this protein alone and in complex with the S100A16 protein will allow a better understanding of the membrane behavior of these proteins,as well as their roles in the maintenance of membrane integrity.
文摘Background:(I)To describe the development and components of the automobile simulator driving behavior evaluation system developed by CRIR-Institut Nazareth et Louis-Braille;(II)to present the preliminary results of the content evaluation of the driving behavior evaluation grid.Methods:The evaluation system consists of five components:(I)the VS500M Car Simulator(Virage Simulation);(II)four VS500M driving scenarios,modified to minimize the occurrence of simulator sickness and expose subjects to commonly encountered driving situations on highways and city boulevards;(III)the Tobii Pro Glasses 2 eye tracking device;(IV)a car simulator driving behavior observation grid(DBOG);(V)a software application used during the behaviour evaluation phase,where synchronized video tracking,certain data from the simulator(e.g.,speed)and the DBOG grid are presented.Initially,the expected safe driving behaviors were identified,including 235 of a visual nature,supported by literature data and consultation of the project steering committee and an expert in driving assessment.Driving behaviors were assessed in 22 subjects without visual impairment(mean age 55±20 years).Subsequently,the items were revised to determine their relevance based on their importance in terms of road safety or on the frequency with which behaviors were observed among participants.For analysis purpose,the items of the DBOG were grouped according to their content,by type of expected driving behavior(e.g.,following a stop,look to the left and right before crossing the intersection)or element to be detected(e.g.,pedestrians).Results:Some visual behaviors are difficult to observe with the eye tracker device because they are more dependent on peripheral than central vision.Others are rarely observed,possibly because they are little or not realized in daily life or the representation of reality on the simulator does not stimulate their adoption.On the other hand,the visual detection behaviors expected in a situation where safety can be compromised are mostly carried out(e.g.,detection of oncoming vehicles,side mirror verification when changing lanes).Conclusions:This first phase of the study has made possible to better target the items to be kept in the car simulator driving behavior observation grid,and those that will have to be removed as they are too difficult to observe or too rarely adopted by the participants.Content validity and inter-rater reliability will be assessed from the simplified grid.
文摘Background:The S100A10 protein might be an early biomarker of diabetes development leading to diabetic retinopathy.The protein complex S100A10/annexin A2 allows the recruitment of the C-terminal of AHNAK protein(AHNAK C-ter peptide)to the membrane in presence of calcium,before forming a platform which can initiate membrane repair.However,no molecular data are currently available on membrane binding of the different proteins involved in this complex.We aim to study the membrane binding of S100A10,AHNAK C-ter peptide and their complex to better understand their roles in cell membrane repair process.Methods:Firstly,S100A10 will be overexpressed and purified by affinity chromatography and AHNAK C-ter peptide will be synthesized.Langmuir monolayers membrane model will then be used to characterize the interactions between these proteins and different phospholipids found in membranes.The secondary structure,orientation and membrane organization of these proteins will be studied by Polarization Modulation Infrared Reflection-Absorption Spectroscopy.Their lateral localization will be determined through the influence of these proteins on the physical state of lipids by fluorescence microscopy.Results:The optimization of the overexpression,purification and cleavage of the GST tag procedure to obtain pure S100A10 was completed.Protein identification by mass spectrometry and circular dichroism stability pre-studies were performed.In parallel,AHNAK C-ter peptide was studied by Langmuir monolayer model and the results indicate this peptide prefers lipids with negatively charged polar heads and unsaturated acyl chains.Preliminary solid-state NMR results confirm this phenomenon at 37℃.Conclusions:Our research will complete current knowledge on membrane binding of S100A10 and AHNAK C-ter peptide.We could also identify the conditions leading to modifications of these membrane bindings,and possibly to the loss of protein function.Thus,this project helps to better determine their roles in membrane repair,as well as in other physiological mechanisms in which these proteins are involved.
基金This study was supported by a grant from the National Natural Science Foundation of China (No. 30470645).
文摘Background The levels of long-term elevated serum or intracellular free fatty acid （FFA） induce insulin resistance associated with central obesity. The insulin-mimetic protein visfatin is preferentially produced by visceral adipose tissues and has been implicated in obesity and insulin resistance. To identify that FFA is capable of inducing insulin resistance and to clarify the role of FFA on visfatin, we examined the effect of monounsaturated FFA oleate （C 18： 1） and saturated FFA palmitate （C 16：0） on glucose transport and visfatin gene expression in cultured 3T3-L1 adipocytes or preadipocytes.Methods FFA-free DMEM/F12, 0.125 mmol/L, 0.5 mmol/1 and 1.0 mmol/L oleate or palmitate was added to cultured 3T3-L1 adipocytes or preadipocytes and incubated overnight. Glucose transport was assessed as 3H- 2-deoxy-glucose uptake. Total RNA was extracted and subjected to RT-PCR for the measurement of visfatin mRNA levels. Statistical comparisons between control group and other groups were performed with the two-tailed paired t test, and one-way ANOVA was used to compare the mean values among the groups.Results Insulin increased specific membrane glucose transport in 3T3-L1 preadipocytes. Upregulation was evident from 15 minutes to 1 hour exposure to insulin. However, after 6-hour exposure to insulin, there was a downregulation in the response to insulin. Dose response studies demonstrated that 2-deoxy glucose transport was increased by 336% at 50 nmol/L insulin （P〈0.01）, and reached a maximal effect at 100 nmol/L insulin （P〈0.01）. Oleate and palmitate treatment did not influence basal glucose transport （without insulin stimulation）, whereas insulin-stimulated glucose transport was inhibited after overnight oleate and palmitate treatment in preadipocytes and adipocytes. In 3T3-L1 preadipocytes, insulin resistance could be achieved at 0.125 mmol/L oleate or palmitate （P〈0.05, respectively）, and the inhibition was dose dependent. In adipocytes, the inhibition was noted at 0.5 mmol/L oleate or 1.0 mmol/L palmitate. Visfatin mRNA expression increased during differentiation more than 1.5-fold. Bovine serum albumin （BSA） did not influence visfatin mRNA expression compared with the control group. Dose-response studies demonstrated that addition of 0.125 mmol/L oleate and palmitate to 3T3-L1 adipocytes decreased visfatin mRNA expression significantly （78%, 77%, respectively, relative to untreated control, P〈0.05）, and further to 65% （relative to untreated control, P〈0.05） and 55% （relative to untreated control, P〈0.01） at 1.0 mmol/L FFA. Furthermore, the suppression on preadipocytes was similar to that of adipocytes, which reached a maximal reduction of 44% （oleate, P〈0.05） and 47% （palmitate, P〈0.05） at 1.0 mmol/L FFA.Conclusions Oleic acid and palmitic acid may induce insulin resistance in 3T3-L1 adipocytes and preadipocytes. Downregulation of visfatin mRNA may contribute to impair insulin sensitivity caused by oleate and palmitate.
文摘Chronic inflammatory lung disease is a common ,health problem and often treated with potent antibiotics, anti-tuberculosis drugs, and antifungal agents. However, in case of medical therapy failure, surgical treatment has been often considered as an effective procedure. Indications for surgical procedure mainly include recurrent infection, acute suppurative complications including lung abscess and empyema, and often chronic, intermittent, or massive hemoptysis.1 Segmentectomy or lobectomy is considered as an ＂ideal＂ procedure when the disease is localized to one lung and single segment or lobe.
