Atorvastatin reduces the expression of aldo-keto reductases in HUVEC and PTEC. A new approach to influence the polyol pathway

Purpose: Increased flux of glucose via the polyol pathway, oxidative stress and ischaemia lead to the upregulation of the aldose reductase (AR), the key enzyme of the polyol pathway. This adversely affects the organism and can in part be reduced by inhibition of the enzyme. Methods: In this study, we examined the effect of the HMG-CoA-reductase inhibitor atorvastatin on the expression of aldose reductase (AR, AKR1B1), aldehyde reductase (AldR, AKR1A1) and small intestine reductase (SIR, AKR1B10) in human umbilical vein endothelial cells (HUVEC) and human proximal tubular epithelial cells (PTEC) by RT-PCR. Results: In HUVEC, atorvastatin reduces the expression of aldehyde reductase and aldose reductase compared with control medium (-20% and -12% respectively, P < 0.05), while small intestine reductase is not expressed. In PTEC no regulation of aldehyde reductase and aldose reductase by atorvastatin could be measured, while the expression of small intestine reductase was reduced by 37% compared with control medium (P < 0.05). The reduction observed was not abolished by the addition of mevalonic acid. Conclusion: The reduction of members of the aldo-keto-reductase family by atorvastatin is a novel way to influence the polyol pathway and a new pleiotropic effect of atorvastatin

Comparison of Fatty Acid and Gene Profiles in Skeletal Muscle in Normal and Obese C57BL/6J Mice before and after Blunt Muscle Injury

Injury and obesity are two major health burdens affecting millions of people worldwide. Obesity is recognized as a state of chronic inflammation accompanied by various co-morbidities like T2D or cardiovascular diseases. There is increasing evidence that obesity impairs muscle regeneration, which is mainly due to chronic inflammation and to excessive accumulation of lipids in adipose and non-adipose tissue. To compare fatty acid profiles and changes in gene expression at different time points after muscle injury, we used an established drop tower-based model with a defined force input to damage the extensor iliotibialis anticus on the left hind limb of female C57BL/6J mice of normal weight and obese mice. Although most changes in fatty acid content in muscle tissue are diet related, levels of eicosaenoic (normal weight) and DHG-linolenic acid (obese) in the phospholipid and docosahexaenoic acid (normal weight) in the triglyceride fraction are altered after injury. Furthermore, changes in gene transcription were detected in 3829 genes in muscles of normal weight mice, whereas only 287 genes were altered in muscles of obese mice after trauma. Alterations were found within several pathways, among them notch-signaling, insulin-signaling, sonic hedgehog-signaling, apoptosis related pathways, fat metabolism related cholesterol homeostasis, fatty acid biosynthetic process, fatty acid elongation, and acyl-CoA metabolic process. We could show that genes involved in fat metabolism are affected 3 days after trauma induction mostly in normal weight but not in obese mice. The strongest effects were observed in normal weight mice for Alox5ap, the activating protein for leukotriene synthesis, and Apobec1, an enzyme substantial for LDL synthesis. In summary, we show that obesity changes the fat content of skeletal muscle and generally shows a negative impact upon blunt muscle injury on various cellular processes, among them fatty acid related metabolism, notch-, insulin-, sonic hedgehog-signaling, and apoptosis

Donor dopamine treatment in brain dead rats is associated with an improvement in renal function early after transplantation and a reduction in renal inflammation

Brain death (BD) is associated with tissue inflammation. As dopamine treatment of BD donor rats reduces renal monocyte infiltration, we tested if this treatment affects renal function and inflammation in recipients. BD was induced in F344 rats and was maintained for 6 h in all experiments. Dopamine was given for 6 (DA6) or 3 h (DA3) from the onset of BD. Ventilated non-BD (NBD) and BD animals served as controls. Kidneys were transplanted into bilaterally nephrectomized Lewis recipients. Serum creatinine (s-crea) was measured and leukocyte infiltration was assessed 10 days after transplantation. One day after transplantation, s-crea was significantly reduced in recipients who received a renal allograft from dopamine treated BD or from NBD rats compared to BD vehicle (P <0.05). Ten days after transplantation, the number of infiltrating monocytes was significantly lower in grafts obtained from dopamine treated and from NBD rats (P <0.05). A reduced infiltration in these grafts was confirmed by Banff 97 classification. Cytokine-induced neutrophil-chemoattractant 1 and interleukin (IL)-6 mRNA expression were reduced in DA rats compared to BD controls. No difference for macrophage chemoattractant protein 1 and IL-10 were found. These findings may explain the salutary effect of donor dopamine treatment in renal transplantation

Table_4_Diet-Induced Obesity Affects Muscle Regeneration After Murine Blunt Muscle Trauma—A Broad Spectrum Analysis.docx

<p>Injury to skeletal muscle affects millions of people worldwide. The underlying regenerative process however, is a very complex mechanism, time-wise highly coordinated, and subdivided in an initial inflammatory, a regenerative and a remodeling phase. Muscle regeneration can be impaired by several factors, among them diet-induced obesity (DIO). In order to evaluate if obesity negatively affects healing processes after trauma, we utilized a blunt injury approach to damage the extensor iliotibialis anticus muscle on the left hind limb of obese and normal weight C57BL/6J without showing any significant differences in force input between normal weight and obese mice. Magnetic resonance imaging (MRI) of the injury and regeneration process revealed edema formation and hemorrhage exudate in muscle tissue of normal weight and obese mice. In addition, morphological analysis of physiological changes revealed tissue necrosis, immune cell infiltration, extracellular matrix (ECM) remodeling, and fibrosis formation in the damaged muscle tissue. Regeneration was delayed in muscles of obese mice, with a higher incidence of fibrosis formation due to hampered expression levels of genes involved in ECM organization. Furthermore, a detailed molecular fingerprint in different stages of muscle regeneration underlined a delay or even lack of a regenerative response to injury in obese mice. A time-lapse heatmap determined 81 differentially expressed genes (DEG) with at least three hits in our model at all-time points, suggesting key candidates with a high impact on muscle regeneration. Pathway analysis of the DEG revealed five pathways with a high confidence level: myeloid leukocyte migration, regulation of tumor necrosis factor production, CD4-positive, alpha-beta T cell differentiation, ECM organization, and toll-like receptor (TLR) signaling. Moreover, changes in complement-, Wnt-, and satellite cell-related genes were found to be impaired in obese animals after trauma. Furthermore, histological satellite cell evaluation showed lower satellite cell numbers in the obese model upon injury. Ankrd1, C3ar1, Ccl8, Mpeg1, and Myog expression levels were also verified by qPCR. In summary, increased fibrosis formation, the reduction of Pax7⁺ satellite cells as well as specific changes in gene expression and signaling pathways could explain the delay of tissue regeneration in obese mice post trauma.</p

Hypothermia-induced loss of endothelial barrier function is restored after dopamine pretreatment:Role of p42/p44 activation

Background. Donor dopamine usage is associated with improved immediate graft function after renal transplantation. Although prolonged cold preservation results in an increased vascular permeability, the present study was conducted to examine in vitro and in vivo if dopamine Pretreatment influences endothelial barrier function under such conditions. Methods. To assess cold preservation injury in vitro and in vivo, cultured human umbilical vein endothelial cells (HUVEC) and Lewis donor rats were pretreated with dopamine or isotonic saline prior to cold storage. Injury was determined by lactate dehydrogenase (LDH) release, histology, and functional analysis. Results. In vitro cold storage resulted in intercellular gap formation in both untreated and dopamine pretreated HUVEC. In the latter monolayer integrity was completely restored upon rewarming and paracellular transport of fluorescein isothiocyanate-dextran was significantly reduced. In dopamine treated HUVEC, intercellular gap formation was independent of cell death and was associated with redistribution of junctional proteins and condensation of cytoskeleton proteins. In untreated HUVEC proteolysis and cell death were clearly evident after hypothermia. Closing of intercellular gaps was dependent on p42/p44 activation. Regeneration of adenosine triphosphate was only observed in dopamine pretreated cells. Only in dopamine treated Lewis renal allografts subjected to cold storage, activation of p42/p44 occurred upon rewarming. These grafts had a better renal function and displayed less inflammatory cells five days after transplantation. Conclusion. Our study demonstrates beneficial effects of dopamine treatment on cold storage induced endothelial barrier disturbances. This may contribute to the positive effects of catecholamines on immediate graft function of renal allografts in men

Table_2_Diet-Induced Obesity Affects Muscle Regeneration After Murine Blunt Muscle Trauma—A Broad Spectrum Analysis.docx

<p>Injury to skeletal muscle affects millions of people worldwide. The underlying regenerative process however, is a very complex mechanism, time-wise highly coordinated, and subdivided in an initial inflammatory, a regenerative and a remodeling phase. Muscle regeneration can be impaired by several factors, among them diet-induced obesity (DIO). In order to evaluate if obesity negatively affects healing processes after trauma, we utilized a blunt injury approach to damage the extensor iliotibialis anticus muscle on the left hind limb of obese and normal weight C57BL/6J without showing any significant differences in force input between normal weight and obese mice. Magnetic resonance imaging (MRI) of the injury and regeneration process revealed edema formation and hemorrhage exudate in muscle tissue of normal weight and obese mice. In addition, morphological analysis of physiological changes revealed tissue necrosis, immune cell infiltration, extracellular matrix (ECM) remodeling, and fibrosis formation in the damaged muscle tissue. Regeneration was delayed in muscles of obese mice, with a higher incidence of fibrosis formation due to hampered expression levels of genes involved in ECM organization. Furthermore, a detailed molecular fingerprint in different stages of muscle regeneration underlined a delay or even lack of a regenerative response to injury in obese mice. A time-lapse heatmap determined 81 differentially expressed genes (DEG) with at least three hits in our model at all-time points, suggesting key candidates with a high impact on muscle regeneration. Pathway analysis of the DEG revealed five pathways with a high confidence level: myeloid leukocyte migration, regulation of tumor necrosis factor production, CD4-positive, alpha-beta T cell differentiation, ECM organization, and toll-like receptor (TLR) signaling. Moreover, changes in complement-, Wnt-, and satellite cell-related genes were found to be impaired in obese animals after trauma. Furthermore, histological satellite cell evaluation showed lower satellite cell numbers in the obese model upon injury. Ankrd1, C3ar1, Ccl8, Mpeg1, and Myog expression levels were also verified by qPCR. In summary, increased fibrosis formation, the reduction of Pax7⁺ satellite cells as well as specific changes in gene expression and signaling pathways could explain the delay of tissue regeneration in obese mice post trauma.</p

Table_1_Diet-Induced Obesity Affects Muscle Regeneration After Murine Blunt Muscle Trauma—A Broad Spectrum Analysis.docx

<p>Injury to skeletal muscle affects millions of people worldwide. The underlying regenerative process however, is a very complex mechanism, time-wise highly coordinated, and subdivided in an initial inflammatory, a regenerative and a remodeling phase. Muscle regeneration can be impaired by several factors, among them diet-induced obesity (DIO). In order to evaluate if obesity negatively affects healing processes after trauma, we utilized a blunt injury approach to damage the extensor iliotibialis anticus muscle on the left hind limb of obese and normal weight C57BL/6J without showing any significant differences in force input between normal weight and obese mice. Magnetic resonance imaging (MRI) of the injury and regeneration process revealed edema formation and hemorrhage exudate in muscle tissue of normal weight and obese mice. In addition, morphological analysis of physiological changes revealed tissue necrosis, immune cell infiltration, extracellular matrix (ECM) remodeling, and fibrosis formation in the damaged muscle tissue. Regeneration was delayed in muscles of obese mice, with a higher incidence of fibrosis formation due to hampered expression levels of genes involved in ECM organization. Furthermore, a detailed molecular fingerprint in different stages of muscle regeneration underlined a delay or even lack of a regenerative response to injury in obese mice. A time-lapse heatmap determined 81 differentially expressed genes (DEG) with at least three hits in our model at all-time points, suggesting key candidates with a high impact on muscle regeneration. Pathway analysis of the DEG revealed five pathways with a high confidence level: myeloid leukocyte migration, regulation of tumor necrosis factor production, CD4-positive, alpha-beta T cell differentiation, ECM organization, and toll-like receptor (TLR) signaling. Moreover, changes in complement-, Wnt-, and satellite cell-related genes were found to be impaired in obese animals after trauma. Furthermore, histological satellite cell evaluation showed lower satellite cell numbers in the obese model upon injury. Ankrd1, C3ar1, Ccl8, Mpeg1, and Myog expression levels were also verified by qPCR. In summary, increased fibrosis formation, the reduction of Pax7⁺ satellite cells as well as specific changes in gene expression and signaling pathways could explain the delay of tissue regeneration in obese mice post trauma.</p