In vitro model for pathological or physiologic conditions

Abstract

The present invention generally relates to in vitro methods for mimicking in vivo pathological or physiologic conditions. The methods comprise applying shear forces to a cell type or cell type plated on a surface within a cell culture container. Methods for testing drugs or compounds in such systems are also described.

1 - 299 . (canceled) 
     
     
         300 . A method of mimicking a pathological or physiologic condition in vitro, the method comprising:
 adding a culture medium and at least one factor to a cell culture container;   plating at least one cell type on at least one surface within the cell culture container, wherein the surface comprises a surface of a porous membrane suspended in the cell culture container; and   applying a shear force upon a second surface of the porous membrane, the shear force resulting from flow of the culture medium induced by a flow device, the flow mimicking flow to which the at least one cell type is exposed in vivo in the pathological or physiologic condition, wherein:   the concentration of the factor in the culture media for mimicking the pathological condition is either:
 (i) within the in vivo concentration range of the factor observed in the pathological condition; or 
 (ii) within the concentration range of the factor that would result in vivo from administration of a drug or a compound; or 
   the concentration of the factor in the culture media for mimicking the physiologic condition is either:
 (i) within the in vivo concentration range of the factor observed in the physiologic condition; or 
 (ii) within the concentration range of the factor that would result in vivo from administration of a drug or a compound. 
   
     
     
         301 . The method of  claim 300 , wherein the method further comprises testing a drug or a compound for an effect on the pathological or physiologic condition, wherein testing the drug or the compound for the effect on the pathological or physiologic condition comprises:
 adding the drug or the compound to the culture medium after plating the at least one cell type; and   applying the shear force upon the at least one cell type exposed to the drug or the compound;   wherein a change in the at least one cell type in the presence of the drug or the compound indicates that the drug or the compound has an effect on the pathological or physiological condition.   
     
     
         302 . The method of  claim 300 , wherein the pathological condition is mimicked. 
     
     
         303 . The method of  claim 302 , wherein the concentration of the factor in the culture media is within the in vivo concentration range of the factor observed in the pathological condition. 
     
     
         304 . The method of  claim 302 , wherein the concentration of the factor in the culture media is within the concentration range of the factor that would result in vivo from administration of a drug or a compound. 
     
     
         305 . The method of  claim 300 , wherein a change in a level of a marker of the pathological condition in the at least one plated cell type or in the culture media upon application of the shear force, as compared to the level of the marker in the at least one plated cell type or in the culture media in the absence of application of the shear force confirms mimicking of the pathological condition. 
     
     
         306 . The method of  claim 300 , wherein the physiologic condition is mimicked. 
     
     
         307 . The method of  claim 306 , wherein the concentration of the factor in the culture media is within the in vivo concentration range of the factor observed in the physiologic condition. 
     
     
         308 . The method of  claim 306 , wherein the concentration of the factor in the culture media is within the concentration range of the factor that would result in vivo from administration of a drug or a compound. 
     
     
         309 . The method of  claim 300 , wherein a change in a level of a marker of the physiologic condition in the at least one plated cell type or in the culture media upon application of the shear force, as compared to the level of the marker in the at least one plated cell type or in the culture media in the absence of application of the shear force confirms mimicking of the physiologic condition. 
     
     
         310 . The method of  claim 300 , wherein the at least one plated cell type comprises renal cells, cells of the airways, blood-brain barrier cells, vascular cells, hepatic cells, pancreatic cells, cardiac cells, muscle cells, spleen cells, gastrointestinal tract cells, skin cells, liver cells, immune cells, or hematopoietic cells. 
     
     
         311 . The method of  claim 300 , wherein the at least one plated cell type comprises astrocytes, endothelial cells, glomerular fenestrated endothelial cells, renal epithelial podocytes, alpha cells, β-cells, delta cells, pancreatic polypeptide (PP) cells, epsilon cells, glial cells, hepatocytes, neurons, nonparenchymal hepatic cells, podocytes, smooth muscle cells, mesangial cells, pericytes, cardiac muscle cells, skeletal muscle cells, leukocytes, monocytes, myocytes, macrophages, neutrophils, dendritic cells, T-cells, B-cells, endothelial progenitor cells, stem cells, circulating stem cells, circulating hematopoietic cells, or a combination of any thereof. 
     
     
         312 . The method of  claim 300 , wherein the pathological condition comprises a vascular pathological condition, the factor comprises oxidized low-density lipoprotein (oxLDL), tumor necrosis factor-α (TNFα), glucose, tissue growth factor-β (TGF-β), an elastin degradation product, elastase, vitamin D, an inorganic phosphate, leptin, adiponectin, apelin, aldosterone, angiotensin II, a triglyceride, high-density lipoprotein (HDL), oxidized high-density lipoprotein (oxHDL), a triglyceride-rich lipoprotein, low-density lipoprotein (LDL), insulin, a fatty acid, or a combination of any thereof. 
     
     
         313 . The method of  claim 312 , wherein the at least one cell type comprises endothelial cells, smooth muscle cells, or endocardial cells. 
     
     
         314 . The method of  claim 300 , wherein the factor comprises TNFα, a fatty acid, or a combination thereof. 
     
     
         315 . The method of  claim 302 , wherein the pathological condition comprises advanced inflammation, atherosclerosis, diabetic nephropathy, diabetic neuropathy, diabetic retinopathy, hypertension, hypertensive encephalopathy, hypertensive retinopathy, fatty liver disease, hypertension, heart failure, stroke, Marfan syndrome, carotid intima-medial thickening, atrial fibrillation, kidney disease, pulmonary fibrosis, chronic obstructive pulmonary disease, hyperlipidemia, hypercholesterolemia, diabetes, atherosclerotic plaque rupture, atherosclerotic plaque erosion, thoracic aortic aneurysm, cerebral aneurysm, abdominal aortic aneurysm, cerebral aneurysm, pulmonary artery disease, pulmonary hypertension, peripheral artery disease, deep vein thrombosis, vascular restenosis, vascular calcification, myocardial infarction, obesity, hypertriglyceridemia, hypoalphalipoproteinemia, hepatitis C, hepatitis B, liver fibrosis, bacterial infection, viral infection, cirrhosis, or alcohol-induced liver disease. 
     
     
         316 . The method of  claim 300 , wherein plating the at least one cell type on the surface of the porous membrane comprises plating a first cell type on a first surface of the porous membrane and plating a second cell type on a second surface of the porous membrane, wherein the shear force is applied upon the second cell type. 
     
     
         317 . The method of  claim 316 , wherein:
 the first cell type comprises smooth muscle cells and the second cell type comprises endothelial cells; or   the first cell type comprises hepatocytes and the second cell type comprises non-parenchymal hepatic cells.   
     
     
         318 . The method of  claim 300 , wherein:
 the porous membrane is positioned in the cell culture container such that a first surface of the porous membrane forms a boundary of a first volume within the container and the second surface forms a boundary of a second volume within the container;   the first volume comprises the at least one cell type; and   the shear force is applied by inducing the flow of the culture medium within the second volume of the container.   
     
     
         319 . An in vitro method of testing a drug or a compound for an effect, the method comprising:
 adding a culture media to a cell culture container;   plating at least one cell type on at least one surface within the cell culture container;   adding a drug or a compound to the culture media, wherein the concentration of the drug or the compound in the culture media is within the concentration range of the drug or the compound that achieves the effect in vivo; and   applying a shear force upon the at least one plated cell type exposed to the drug or the compound, the shear force resulting from flow of the culture media induced by a flow device, the flow mimicking flow to which the at least one cell type is exposed in vivo, wherein a change in the at least one plated cell type, in the presence of the drug or the compound, indicates that the drug or the compound has the effect.