Dr. László Nagy, Dr. Bálint László Bálint, Dr. Bálint L. Bálint L., Dr. Bertalan Meskó, Dr. László Nagy, Dr. Árpád Lányí, Dr. Beáta Scholtz, Dr. Lajos Széles, Dr. Tamás Varga (2011)
University of Debrecen
The transcription factor PPARgamma is involved the differentiation of adipocytes and well- established cellular models are available to study this differentiation pathways. With these models, high throughput screens can be performed, where hundreds or even thousands of ligands can be screened to assess their role in adipose tissue differentiation, whereby one can estimate if these ligands as drugs cause weight gain or not as a side effect of the adipose differentiation.
The molecular mechanisms of adipocyte differentiation
The PPAR gamma is a well-characterized transcription factor and its role in adipocyte differentiation is well-known. There are established cellmodels of adipocyte differentiation, such as 3T3L1 mouse fibroblasts or embryonic stem cells. Immature adipocytes can be differentiated from fibroblasts adipoblasts, preadipocyteadipocyte by an endpoint of differentiation in the adipocyteadipocyte, which is filled with lipid droplets. In such a cell model several tens, hundreds or even hundreds of thousands of ligands can be tested in order to see if they influence adipocyte adipocyte differentiation, and thereby to establish if they increase weight if administered as a drug.
PPARgamma is modulating the metabolism through several tissues. Since PPARgamma is a transcription factor that forms heterodimers with RXR, it can directly cause changes in gene expression. The three main tissues where it is influencing metabolism are the hepatic, the adipose tissues and the muscle. Other effects of PPARgamma include changes in the tolerogenic activity of immune cells. Synthetic PPAR gamma ligands are administered in type two diabetes due to the fact that they reduce the insulin resistance of the peripheral tissues, mainly the muscles. Another effect of them is the facilitation of terminal differentiation of adipocytes. This effect is producing weight gain in the subcutaneous adipose tissues and redistribution from visceral fat to subcutaneous fat.
Through CD36 and ap2 or FABP4 it is increasing the production of triglycerides. In hepatocytes PPARgamma is increasing beta oxidation in mitochondria and peroxisomes.
PPARg and lipid uptake
Cholesterol is transported in the circulation in the form of lipoprotein complexes. LDL is able to leave the circulation and is responsible for the cholesterol supply of the peripheral cells through the LDL receptors. In some of the cases, the lipids of the LDL are present in an oxidized form, because of smoking or because of the fried oils of dietary origin. In these cases, the LDL is forming oxidized LDL, which is taken up by scavenger receptors and by CD36. Oxidized LDL contains PPARg ligands, which, if activated, will further increase the production of CD36, whereby a self upregulated loop is initiated, which will lead to the filling of the macrophages with cholesterol, the formation of foamy cells and the death of the macrophage with a local inflammation. The result is the formation of the atherosclerotic plaque.
There is a nuclear receptor driven pathway that is able to export the cholesterol from the cell. The PPARg agonists from the oxidized LDL are turning on the expression of LXR. If LXR ligands are present, LXR will turn on the synthesis of ABCA1 the main ATP driven cholesterol pump of the cells. As a result, cholesterol is pumped into the HDL molecules and transported back to the liver.
LXR will turn on the synthesis of ABCA1 the main ATP driven cholesterol pump of the cells. As a result cholesterol is pumped into the HDL molecules and transported back to the liver.
Cholesterol has a very low water solubility, in the circulation it is transported in the form of lipoprotein complexes. Some of the lipoprotein complexes are quite large and involved mainly in intravascular transport, like chlyomicrons and VLDL other, smaller complexes like LDL and HDL can leave the circulation and are involved in direct transfer from and to the cells. Chylomicrons can be conceived as huge lipid droplets that leave the enterocyte and arrive into the lymph. After initial processing, they get into the circulation and the chylomicron remnants are taken up by the hepatocytes. In the hepatocytes, the lipids are repacked into VLDL. VLDL is transformed into IDL and LDL in the circulation and LDL leaves the circulation. HDL is produced mainly in hepatocytes and to a lesser extent in enterocytes. As an empty complex arrives at the periphery where it is filled with cholesterol by the ATP dependent transporter ABCA1. The function of HDL is the transport of retrograde cholesterol from the periphery to the liver, the supply of cholesterol of the endocrine organs and the moving of lipoproteins between the lipoprotein complexes in the circulation. Bile acids produced in the liver arrive at the intestine with the bile and they are involved in the emulsification of lipids in the intestine and thus in the uptake of the lipids in the intestine.
Uptake of cholesterol into the cells
Cholesterol is taken up by the cells in the form of LDL. The highest number of LDL receptors are on the hepatocytes and the expression of LDL receptors is controlled by the level of cholesterol in the cells through the cholesterol sensing molecule SREBP1. The first step of the uptake is the binding of LDL by the LDL receptors expressed on the cell surface. Mutations in the LDL receptors are causing an increased level of circulating cholesterol. In the case of homozygotic form of the mutation, the patients will have extremely high levels of cholesterol and atherosclerosis at a very young age, which can manifest itself even as myocardial heart infarction. In this case, the single chance to prolong the patient’s life at present is liver transplantation. The new liver will have healthy LDL receptors whereby it will be able to remove the LDL from circulation.
The LDL is taken up through coated pits and it arrives in lysosomes where the proteins are hydrolyzed and the cholesterol enters the cell membranes. LDL receptors recirculate every 10-20 minutes. Their half-life is in the range of 10 to 30 hours and through their life cycle they are involved in the uptake of hundreds of cholesterol molecules.
Through this property of theirs, they can play a role in regenerative medicine, but, more importantly, through this feature we can have insight into the details of differentiation pathways. This knowledge can be used for the differentiation of the induced pluripotent cells (iPS) as well. iPS cells have two other very important proterties. First: they are immunologically identical with the organism which they are originating from and through this they stand a good chance of being used in regenerative medicine applications.
Second: because they are genetically identical with the organism they originate from, they can serve as a source of cellular models of these genetic diseases, whereby we can gain an insight into the molecular details of these diseases.
In the figure, we can see the differentiation steps of embryonic stem cells. In the first step ES cells generate mesenchymal stem cells under the controll of BMP and FGF signals. Mesenchymal stem cells are able to differentiate into myoblasts, adipoblastst and osteoblasts. From this step on, tissue specific transcription factors such as MyoD+Myogenin, PPARg+CEBPa, or RunX2+Osx are involved in the terminal differentiation of the previously mentioned cell types.
Embryonic stem cells are able to differentiate along all differentiating pathways