sirolimus and Atrophy

There is no evidence on the incidence of subclinical inflammation and scaring lesions in patients receiving tacrolimus (TAC) minimization and elimination immunosuppressive regimens.. This study analyzed preimplantation, 3 and 24 months protocol biopsies and anti-HLA donor-specific antibodies (DSA) in 140 low immunological risk kidney transplant recipients receiving reduced TAC exposure, prednisone, and mycophenolate, randomized at 3 months to be converted or not to sirolimus (SRL).. Mean TAC concentrations were 6.0 ± 2.4 ng/mL and 5.8 ± 2.2 ng/mL at 3 and 24 months. The incidence of subclinical inflammation lesions at 3 months was 9.3%. The incidence of (interstitial fibrosis) IF/(tubular atrophy) TA at month 24 was 57.6%, higher in SRL compared to TAC group (68.8 vs 44.4%; P = 0.022). Patients converted to SRL showed higher incidence of acute rejection (7.3% vs 0%), proteinuria (59.6% vs 25%; P = 0.001), and DSA (17.8% vs 7.3%; P = 0.201), respectively. Biopsy-proven acute rejection (odds ratio [OR] 2.32, 95% confidence interval [95% CI], 0.979-5.518, P = 0.056), subclinical inflammation lesions at 3 months (OR, 11.75; 95% CI, 1.286-107.474; P = 0.029) and conversion to SRL (OR, 2.72; 95% CI, 1.155-6.383; P = 0.022) were associated with IF/TA at month 24. Black ethnicity (OR, 0.22; 95% CI, 0.058-0.873; P = 0.031), donor age (OR, 2.74; 95% CI, 1.329-5.649; P = 0.006), and conversion to SRL (OR, 2.34; 95% CI, 1.043-5.267; P = 0.039) were associated with inferior renal function at 24 months.. In kidney transplant recipients receiving reduced TAC exposure, subclinical inflammation lesions at 3 months were associated with IF/TA at 24 months. Conversion from TAC to SRL was associated with inferior renal function, higher incidence of IF/TA, and trends to higher incidence of DSA at 24 months.

Topics: Adult; Atrophy; Biomarkers; Biopsy; Calcineurin Inhibitors; Chi-Square Distribution; Drug Substitution; Female; Fibrosis; Graft Rejection; Graft Survival; Histocompatibility; HLA Antigens; Humans; Immunosuppressive Agents; Isoantibodies; Kidney; Kidney Transplantation; Logistic Models; Male; Middle Aged; Multivariate Analysis; Nephritis; Odds Ratio; Proteinuria; Risk Factors; Sirolimus; Tacrolimus; Time Factors; Treatment Outcome

Calcineurin-inhibitor therapy is a contributing factor to the origin of interstitial fibrosis and tubular atrophy (IFTA).. We conducted a prospective randomized trial of conversion of tacrolimus to sirolimus at 1-month posttransplant in kidney transplant recipients on rapid steroid withdrawal. We compared the chronic changes (IFTA and sum of Banff chronic scores--Total Score) on protocol biopsies at 1 month, 1 year, and 2 years in all randomized patients. We compared the outcomes between treatment groups and analyzed the impact of previous rejection on the chronic changes.. We randomized 122 patients, 62 to sirolimus and 60 to tacrolimus. The 1-year biopsy was performed in 54 patients (90%) of the tacrolimus group and 56 patients (90%) of the sirolimus group. The proportion of biopsies with IFTA more than or equal to 2 and the Total Score more than 2 increased over the 2 years but were not different between the study groups at any time point. On the 1-year biopsy, there was more IFTA, and the fraction with Total Score more than 2 was higher in the tacrolimus group with previous rejection. In the cohort without rejection, there was a significant progression of the IFTA and Total Score between 1 and 2 years in both the sirolimus and tacrolimus groups.. Conversion from tacrolimus to sirolimus at 1-month posttransplant in kidney transplant recipients on rapid steroid withdrawal does not decrease the progression of chronic changes on protocol biopsies during the first 2 years even in those patients without previous acute rejection.

Topics: Adult; Aged; Aged, 80 and over; Atrophy; Biopsy; Contraindications; Disease Progression; Female; Fibrosis; Graft Rejection; Humans; Immunosuppressive Agents; Kidney; Kidney Transplantation; Longitudinal Studies; Male; Middle Aged; Prospective Studies; Risk Factors; Sirolimus; Steroids; Tacrolimus; Transplantation, Homologous; Withholding Treatment

Interstitial fibrosis and tubular atrophy, a major cause of kidney allograft dysfunction, has been linked to premature cellular senescence. The mTOR inhibitor Rapamycin protects from senescence in experimental models, but its antiproliferative properties have raised concern early after transplantation particularly at higher doses. Its effect on senescence has not been studied in kidney transplantation, yet. Rapamycin was applied to a rat kidney transplantation model (3 mg/kg bodyweight loading dose, 1.5 mg/kg bodyweight daily dose) for 7 days. Low Rapamycin trough levels (2.1-6.8 ng/mL) prevented the accumulation of p16INK4a positive cells in tubules, interstitium, and glomerula. Expression of the cytokines MCP-1, IL-1β, and TNF-α, defining the proinflammatory senescence-associated secretory phenotype, was abrogated. Infiltration with monocytes/macrophages and CD8+ T-lymphocytes was reduced and tubular function was preserved by Rapamycin. Inhibition of mTOR was not associated with impaired structural recovery, higher glucose levels, or weight loss. mTOR inhibition with low-dose Rapamycin in the immediate posttransplant period protected from premature cellular senescence without negative effects on structural and functional recovery from preservation/reperfusion damage, glucose homeostasis, and growth in a rat kidney transplantation model. Reduced senescence might maintain the renal regenerative capacity rendering resilience to future injuries resulting in protection from interstitial fibrosis and tubular atrophy.

Topics: Animals; Atrophy; Cellular Senescence; Female; Fibrosis; Glucose; Humans; Kidney; Kidney Diseases; Kidney Transplantation; Male; Rats; Sirolimus; TOR Serine-Threonine Kinases

Glucocorticoids have excellent therapeutic properties; however, they cause significant adverse atrophogenic effects. The mTORC1 inhibitor REDD1 has been recently identified as a key mediator of glucocorticoid-induced atrophy. We performed computational screening of a connectivity map database to identify putative REDD1 inhibitors. The top selected candidates included rapamycin, which was unexpected because it inhibits pro-proliferative mTOR signaling. Indeed, rapamycin inhibited REDD1 induction by glucocorticoids dexamethasone, clobetasol propionate, and fluocinolone acetonide in keratinocytes, lymphoid cells, and mouse skin. We also showed blunting of glucocorticoid-induced REDD1 induction by either catalytic inhibitor of mTORC1/2 (OSI-027) or genetic inhibition of mTORC1, highlighting role of mTOR in glucocorticoid receptor signaling. Moreover, rapamycin inhibited glucocorticoid receptor phosphorylation, nuclear translocation, and loading on glucocorticoid-responsive elements in REDD1 promoter. Using microarrays, we quantified a global effect of rapamycin on gene expression regulation by fluocinolone acetonide in human keratinocytes. Rapamycin inhibited activation of glucocorticoid receptor target genes yet enhanced the repression of pro-proliferative and proinflammatory genes. Remarkably, rapamycin protected skin against glucocorticoid-induced atrophy but had no effect on the glucocorticoid anti-inflammatory activity in different in vivo models, suggesting the clinical potential of combining rapamycin with glucocorticoids for the treatment of inflammatory diseases.

Topics: Animals; Atrophy; Disease Models, Animal; Female; Immunosuppressive Agents; Keratinocytes; Mice; Mice, Inbred C57BL; Receptors, Glucocorticoid; Sirolimus; Skin; Steroids; Transcription Factors

Transit amplifying cells (TACs) are highly proliferative in nature and tend to be sensitive to ionizing radiation. Due to the abundance of TACs that support the elongation of hair shafts, growing hair follicles are highly sensitive to radiation injury. How hair follicles repair themselves after radiation injury is unclear. In this study, we observed that in 4 Gy irradiated mice, hair follicle dystrophy was induced with apoptosis-driven loss of hair matrix cells, which are the TACs that fuel hair growth. The dystrophy was repaired within 96 h without significant hair loss, indicating that a regenerative attempt successfully restored the TAC population to resume anagen growth. Soon after irradiation, mTORC1 signaling was activated in the TAC compartment and its activation was maintained until the regeneration process was completed. Inhibition of mTORC1 by rapamycin treatment increased radiation-induced cell apoptosis, reduced cell proliferation and delayed restoration of Wnt signaling in the hair matrix after radiation injury, leading to prolonged dystrophy and hair loss. These results demonstrate that mTORC1 signaling is activated after irradiation and is required for timely regeneration of the TAC pool of hair follicles, so that hair growth can resume after radiation injury.

Topics: Alopecia; Animals; Apoptosis; Atrophy; Female; Hair; Hair Follicle; Mechanistic Target of Rapamycin Complex 1; Mice; Radiation Injuries, Experimental; Regeneration; Signal Transduction; Sirolimus; Wnt Signaling Pathway

Mechanical ventilation (MV) is associated with atrophy and weakness of the diaphragm muscle, a condition termed ventilator-induced diaphragmatic dysfunction (VIDD). Autophagy is a lysosomally mediated proteolytic process that can be activated by oxidative stress, which has the potential to either mitigate or exacerbate VIDD. The primary goals of this study were to (1) determine the effects of MV on autophagy in the diaphragm and (2) evaluate the impact of antioxidant therapy on autophagy induction and MV-induced diaphragmatic weakness.. Mice were assigned to control (CTRL), MV (for 6 h), MV + N-acetylcysteine, MV + rapamycin, and prolonged (48 h) fasting groups. Autophagy was monitored by quantifying (1) autophagic vesicles by transmission electron microscopy, (2) messenger RNA levels of autophagy-related genes, and (3) the autophagosome marker protein LC3B-II, with and without administration of colchicine to calculate the indices of relative autophagosome formation and degradation. Force production by mouse diaphragms was determined ex vivo.. Diaphragms exhibited a 2.2-fold (95% CI, 1.8 to 2.5) increase in autophagic vesicles visualized by transmission electron microscopy relative to CTRL after 6 h of MV (n = 5 per group). The autophagosome formation index increased in the diaphragm alone (1.5-fold; 95% CI, 1.3 to 1.8; n = 8 per group) during MV, whereas prolonged fasting induced autophagosome formation in both the diaphragm (2.5-fold; 95% CI, 2.2 to 2.8) and the limb muscle (4.1-fold; 95% CI, 1.8 to 6.5). The antioxidant N-acetylcysteine further augmented the autophagosome formation in the diaphragm during MV (1.4-fold; 95% CI, 1.2 to 1.5; n = 8 per group) and prevented MV-induced diaphragmatic weakness. Treatment with the autophagy-inducing agent rapamycin also largely prevented the diaphragmatic force loss associated with MV (n = 6 per group).. In this model of VIDD, autophagy is induced by MV but is not responsible for diaphragmatic weakness. The authors propose that autophagy may instead be a beneficial adaptive response that can potentially be exploited for therapy of VIDD.

Topics: Animals; Antioxidants; Atrophy; Autophagy; Cystine; Diaphragm; Male; Mice; Mice, Inbred C57BL; Muscle Contraction; Phagosomes; Sirolimus; Ventilator-Induced Lung Injury

Salivary gland atrophy is a frequent consequence of head and neck cancer irradiation therapy but can potentially be regulated through the mammalian target of rapamycin (mTOR). Excretory duct ligation of the mouse submandibular gland provokes severe glandular atrophy causing activation of mTOR. This study aims to discover the effects of blocking mTOR signaling in ligation-induced atrophic salivary glands. Following 1 week of unilateral submandibular excretory duct ligation: gland weights were significantly reduced, 4E-BP1 and S6rp were activated, and tissue morphology revealed typical signs of atrophy. However, 3 days following ligation with rapamycin treatment, a selective mTOR inhibitor, gland weights were maintained, 4E-BP1 and S6rp phosphorylation was inhibited, and there were morphological signs of recovery from atrophy. However, following 5 and 7 days of ligation and rapamycin treatment, glands expressed active mTOR and showed signs of considerable atrophy. This evidence suggests that inhibition of mTOR by rapamycin delays ligation-induced atrophy of salivary glands.

Topics: Acinar Cells; Animals; Atrophy; Female; Immunoblotting; Ligation; Mice, Inbred ICR; Organ Size; Phosphorylation; Ribosomal Protein S6; Salivary Ducts; Sirolimus; Staining and Labeling; Submandibular Gland; Time Factors; TOR Serine-Threonine Kinases

Cell cycle arrest is not yet senescence. When the cell cycle is arrested, an inappropriate growth-promotion converts an arrest into senescence (geroconversion). By inhibiting the growth-promoting mTOR pathway, rapamycin decelerates geroconversion of the arrested cells. And as a striking example, while causing arrest, p53 may decelerate or suppress geroconversion (in some conditions). Here I discuss the meaning of geroconversion and also the terms gerogenes, gerossuppressors, gerosuppressants, gerogenic pathways, gero-promoters, hyperfunction and feedback resistance, regenerative potential, hypertrophy and secondary atrophy, pro-gerogenic and gerogenic cells.

Topics: Animals; Atrophy; Cell Cycle Checkpoints; Cellular Senescence; Feedback, Physiological; Humans; Hypertrophy; Protein Kinase Inhibitors; Signal Transduction; Sirolimus; Terminology as Topic; TOR Serine-Threonine Kinases; Tumor Suppressor Protein p53

Chronic renal transplant dysfunction is histopathologically characterized by interstitial fibrosis and tubular atrophy. This study investigated the relative contribution of baseline donor, recipient, and transplant characteristics to interstitial fibrosis and tubular atrophy score at month 12 after renal transplantation.. This retrospective study includes all 109 consecutive recipients with adequate implantation and month 12 biopsies transplanted between April of 2003 and February of 2007. Immunosuppression regimen was tacrolimus and steroids (10 days) plus either sirolimus or mycophenolate mofetil.. Average interstitial fibrosis and tubular atrophy score increased from 0.70 to 1.65 (P<0.001). In an adjusted multiple linear regression analysis, interstitial fibrosis and tubular atrophy score at month 12 was significantly related to donor type (donors after cardiac death versus living donor had interstitial fibrosis and tubular atrophy score+0.41, 95% confidence interval=0.05-0.76, P=0.02), baseline interstitial fibrosis and tubular atrophy, and immunosuppression regimen. Because of interaction between the latter two variables (P=0.002), results are given separately: recipients with a baseline interstitial fibrosis and tubular atrophy score of zero had a 0.60 higher score at month 12 (95% confidence interval=0.09-1.10, P=0.02) when mycophenolate mofetil-treated, whereas recipients with a baseline interstitial fibrosis and tubular atrophy score more than zero had a 0.38 higher score at month 12 (95% confidence interval=0.01-0.74, P=0.04) when sirolimus-treated. A higher score at month 12 correlated with a lower estimated GFR (ρ=-0.45, P<0.001).. These findings suggest that histologic assessment of a preimplantation biopsy may guide choice of immunosuppresion to maximize transplant survival and its interaction with type of immunosuppression.

Topics: Adult; Aged; Atrophy; Biopsy; Drug Therapy, Combination; Female; Fibrosis; Graft Rejection; Graft Survival; Humans; Immunosuppressive Agents; Kidney Transplantation; Kidney Tubules; Linear Models; Male; Middle Aged; Multivariate Analysis; Mycophenolic Acid; Netherlands; Proteinuria; Retrospective Studies; Risk Assessment; Risk Factors; Sirolimus; Steroids; Tacrolimus; Time Factors; Treatment Outcome

Although it is well established that chronic hypoxia leads to an inexorable loss of skeletal muscle mass in healthy subjects, the underlying molecular mechanisms involved in this process are currently unknown. Skeletal muscle atrophy is also an important systemic consequence of chronic obstructive pulmonary disease (COPD), but the role of hypoxemia in this regulation is still debated. Our general aim was to determine the molecular mechanisms involved in the regulation of skeletal muscle mass after exposure to chronic hypoxia and to test the biological relevance of our findings into the clinical context of COPD. Expression of positive and negative regulators of skeletal muscle mass were explored 1) in the soleus muscle of rats exposed to severe hypoxia (6,300 m) for 3 wk and 2) in vastus lateralis muscle of nonhypoxemic and hypoxemic COPD patients. In rodents, we observed a marked inhibition of the mammalian target of rapamycin (mTOR) pathway together with a strong increase in regulated in development and DNA damage response 1 (REDD1) expression and in its association with 14-3-3, a mechanism known to downregulate the mTOR pathway. Importantly, REDD1 overexpression in vivo was sufficient to cause skeletal muscle fiber atrophy in normoxia. Finally, the comparative analysis of skeletal muscle in hypoxemic vs. nonhypoxemic COPD patients confirms that hypoxia causes an inhibition of the mTOR signaling pathway. We thus identify REDD1 as a negative regulator of skeletal muscle mass during chronic hypoxia. Translation of this fundamental knowledge into the clinical investigation of COPD shows the interest to develop therapeutic strategies aimed at inhibiting REDD1.

Topics: Animals; Atrophy; Down-Regulation; Humans; Hypoxia; Male; Mammals; Muscle, Skeletal; Muscular Atrophy; Proto-Oncogene Proteins c-akt; Pulmonary Disease, Chronic Obstructive; Rats; Rats, Wistar; Signal Transduction; Sirolimus

Dietary protein deficiency results in diminished capacity of the pancreas to secrete enzymes needed for macronutrient digestion. Previous work has suggested that modulation of the mammalian target of rapamycin (mTOR) pathway by the hormone cholecystokinin (CCK) plays an important role in normal digestive enzyme synthesis after feeding. The purpose of this study was to elucidate the role of mTOR in protein deficiency-induced pancreatic dysfunction.. Wild-type and CCK-null mice were fed protein-deficient chow for 4 days and then allowed to recover on control chow in the presence or absence of the mTOR inhibitor rapamycin.. The size and secretory capacity of the pancreas rapidly decreased after feeding protein-deficient chow. Refeeding protein-replete chow reversed these changes in both wild-type and CCK-null mice. Changes in the size of the pancreas were paralleled by changes in the content and secretion of digestive enzymes, as well as the phosphorylation of downstream targets of mTOR. Administration of the mTOR inhibitor rapamycin decreased regrowth of the pancreas but did not affect digestive enzyme content or secretory capacity.. These studies demonstrate that dietary protein modulates pancreatic growth, but not digestive enzyme synthesis, via CCK-independent activation of the mTOR pathway.

Topics: Animals; Atrophy; Carrier Proteins; Cholecystokinin; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Organ Size; Pancreas; Phosphotransferases (Alcohol Group Acceptor); Protein Deficiency; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

We investigated the mechanism by which B lymphocyte stimulator (BLyS)/BAFF, a tumor necrosis factor superfamily ligand, promotes B-cell survival and resistance to atrophy. BLyS stimulation activates 2 independent signaling pathways, Akt/mTOR and Pim 2, associated with cell growth and survival. BLyS blocks the cell volume loss (atrophy) that freshly isolated B cells normally undergo when maintained in vitro while concurrently increasing glycolytic activity and overall metabolism. This atrophy resistance requires Akt/mTOR. We used a genetic approach to resolve the contributions of Akt/mTOR and Pim kinase pathways to BLyS-mediated survival. Pim 2-deficient B cells are readily protected from death by BLyS stimulation, but this protection is completely abrogated by treatment with the mTOR inhibitor rapamycin. Furthermore, rapamycin treatment in vivo significantly reduces both follicular and marginal zone B cells in Pim-deficient but not healthy hosts. BLyS-dependent survival requires the antiapoptotic protein Mcl-1. Mcl-1 protein levels rise and fall in response to BLyS addition and withdrawal, respectively, and conditional deletion of the Mcl-1 gene renders B cells refractory to BLyS-mediated protection. Because BlyS is required for the normal homeostasis of all B cells, these data suggest a therapeutic strategy simultaneously inhibiting mTOR and Pim 2 could target pathogenic B cells.

Topics: Animals; Atrophy; B-Cell Activating Factor; B-Lymphocytes; Cell Death; Cell Size; Cell Survival; Germinal Center; Glycolysis; Immunosuppressive Agents; Mice; Mice, Knockout; Myeloid Cell Leukemia Sequence 1 Protein; Neoplasm Proteins; Protein Kinases; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

RB1-inducible Coiled-Coil 1 (RB1CC1) has been shown to be a novel tumor suppressor regulating RB1 expression. Neuronal abundance of RB1CC1 is reported to contribute to the non-proliferating enlarged cell phenotype through the maintenance of RB1 and mTOR. To clarify whether RB1CC1 insufficiency is involved in neuronal atrophy and Alzheimer's pathology, we investigated modifications of RB1CC1 as a possible cause of atrophy or death through the disturbance of mTOR signaling in Neuro-2a neuroblastoma cells. We also evaluated the correlation between RB1CC1 and mTOR signaling in a series of Alzheimer's brain tissues. Though RB1CC1 introduction enhanced neurite growth, RNAi-mediated knockdown of RB1CC1 or rapamycin treatment caused neurite atrophy and apoptosis due to mTOR signaling reduction in the differentiated Neuro-2a cells. Both TSC1 and RB1CC1 were equally functional and maintained mTOR signaling, indicated by phospho-S6 (Ser240/244) expression in 69% of Alzheimer's (9/13 cases) and 100% of normal brains (6/6 cases). However, scanty RB1CC1 expression, less than TSC1, caused phospho-S6 disappearance in 31% of Alzheimer's tissues (4/13 cases). These findings suggest that RB1CC1 insufficiency may result in mTOR signaling repression through unbalanced TSC1 abundance and may induce neuronal atrophy. These observations may have implications for the pathogenesis of Alzheimer's disease.

Topics: Alzheimer Disease; Animals; Atrophy; Autophagy-Related Proteins; Brain; Cell Cycle; Cell Death; Cell Line, Tumor; Embryo, Mammalian; Humans; Immunosuppressive Agents; Intracellular Signaling Peptides and Proteins; Mice; Neuroblastoma; Neurons; Protein Kinases; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Transfection; Tuberous Sclerosis Complex 1 Protein; Tumor Suppressor Proteins

We have previously shown that the common feature of both pressure overload-induced hypertrophy and atrophy is a reactivation of the fetal gene program. Although gene expression profiles and signal transduction pathways in pressure overload hypertrophy have been well studied, little is known about the mechanisms underlying atrophic remodeling of the unloaded heart. Here, we induced atrophic remodeling by heterotopic transplantation of the rat heart. The activity parameters of three signal transduction pathways important in hypertrophy, i.e. mitogen-activated protein (MAP) kinase, mammalian target of rapamycin (mTOR), and Janus kinase/signal transducers and activators of transcription (JAK/STAT), were interrogated. Gene expression of upstream stimuli--insulin-like growth factor 1 (IGF-1) and fibroblast growth factor 2 (FGF-2)--and metabolic correlates, i.e. peroxisome proliferator-activated receptor-alpha (PPARalpha) and PPARalpha-regulated genes, of these pathways were also measured. In addition, we measured transcript levels of genes known to regulate skeletal muscle atrophy, all of which are negatively regulated by IGF-1 (Mafbx/Atrogin-1, MuRF-1). Atrophic remodeling of the heart was associated with increased expression of IGF-1 and FGF-2. Transcript levels of the nuclear receptor PPARalpha were decreased, as were the levels of PPARalpha-regulated genes. Furthermore, there was phosphorylation of ERK1, STAT3, and p70S6K with unloading. Consistent with the increase in IGF-1, we found a decrease in Mafbx/Atrogin-1 and MuRF-1 transcript levels. Rapamycin administration at 0.8 mg/kg/day for 7 days resulted in enhanced atrophy and attenuated the phosphorylation of ERK1, STAT3, and p70S6K without altering gene expression. We conclude that there is significant crosstalk between the mTOR, MAP kinase, and JAK/STAT signaling cascades. Furthermore, ubiquitin ligases, known to be essential for skeletal muscle atrophy, decrease in unloading-induced cardiac atrophy.

Topics: Animals; Atrophy; Fibroblast Growth Factor 2; Gene Expression Profiling; Heart; Heart Transplantation; Insulin-Like Growth Factor I; Male; Mitogen-Activated Protein Kinases; Myocardium; PPAR alpha; Rats; Rats, Wistar; Sirolimus; Ubiquitin-Protein Ligases; Ventricular Remodeling

Rapamycin (Rapa) is an immunosuppressant that is used in patients and animal models to control allograft rejection. Its mechanisms of action are not fully understood. In this article, the authors have investigated the effects of therapeutic doses of Rapa on both thymic and peripheral T-cell populations in the adult rat.. The therapeutic dosage of Rapa was optimized using cardiac transplantation between LEW and DA rats. Thymic morphology was assessed by hematoxylin-eosin staining. Flow cytometric analysis was performed to analyze T-cell phenotype and apoptosis. T-cell receptor (TCR)-mediated T-cell responsiveness was evaluated by 3[H]-thymidine deoxyribose incorporation.. Rapa induced atrophy in the thymus but not in peripheral lymphoid organs. Moreover, fibrosis occurred in thymus that was long-lasting after Rapa withdrawal. In animals treated with Rapa, there was a significant reduction in CD4+CD8+ thymocytes caused by accelerated apoptosis, whereas CD4-CD8-, CD4+CD8-, and CD8+CD4- populations remained unaffected. In contrast, the cellularity of the periphery lymphoid organs was not altered. Within the CD4+ thymocyte population, CD4+CD25+ thymocytes were resistant to Rapa-accelerated apoptosis, and in the periphery, the ratio of CD4+CD25+ to CD4+CD25- T cells was increased. Notably, the peripheral CD4+CD25+ T cells were hyporesponsive to TCR-mediated activation.. The resistance of the peripheral CD4+CD25+ T cells to Rapa treatment might contribute to its immunosuppressive action. The long-term effects of Rapa on thymus atrophy and thymocyte development requires consideration with respect to its clinical application.

Topics: Animals; Apoptosis; Atrophy; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Heart Transplantation; Immunosuppressive Agents; Lymphocyte Activation; Male; Models, Animal; Rats; Rats, Inbred Lew; Receptors, Interleukin-2; Sirolimus; T-Lymphocyte Subsets; T-Lymphocytes; Thymus Gland; Transplantation, Homologous

Mechanical unloading of the heart results in atrophic remodeling. In skeletal muscle, atrophy is associated with inactivation of the mammalian target of rapamycin (mTOR) pathway and upregulation of critical components of the ubiquitin proteosome proteolytic (UPP) pathway. The hypothesis is that mechanical unloading of the mammalian heart has differential effects on pathways of protein synthesis and degradation.. In a model of atrophic remodeling induced by heterotopic transplantation of the rat heart, we measured gene transcription, protein expression, polyubiquitin content, and regulators of the mTOR pathway at 2, 4, 7, and 28 days. In atrophic hearts, there was an increase in polyubiquitin content that peaked at 7 days and decreased by 28 days. Furthermore, gene and protein expression of UbcH2, a ubiquitin conjugating enzyme, was also increased early in the course of unloading. Transcript levels of TNF-alpha, a known regulator of UbcH2-dependent ubiquitin conjugating activity, were upregulated early and transiently in the atrophying rat heart. Unexpectedly, p70S6K and 4EBP1, downstream components of mTOR, were activated in atrophic rat heart. This activation was independent of Akt, a known upstream regulator of mTOR. Rapamycin treatment of the unloaded rat hearts inhibited the activation of p70S6K and 4EBP1 and subsequently augmented atrophy in these hearts compared with vehicle-treated, unloaded hearts.. Atrophy of the heart, secondary to mechanical unloading, is associated with early activation of the UPP. The simultaneous activation of the mTOR pathway suggests active remodeling, involving both protein synthesis and degradation.

Topics: Animals; Atrophy; Cardiomyopathies; Disease Models, Animal; Male; Organ Size; Phosphorylation; Protein Kinase Inhibitors; Protein Kinases; Proteins; Rats; Rats, Wistar; RNA, Messenger; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Transplantation, Heterotopic; Tumor Necrosis Factor-alpha; Ubiquitin-Conjugating Enzymes; Ventricular Remodeling

TOR is a phosphatidylinositol kinase (PIK)-related kinase that controls cell growth and proliferation in response to nutritional cues. We describe a C. elegans TOR homolog (CeTOR) and phenotypes associated with CeTOR deficiency. These phenotypes are compared with the response to starvation and the inactivation of a variety of putative TOR targets.. Whether caused by mutation or RNA interference, TOR deficiency results in developmental arrest at mid-to-late L3, which is accompanied by marked gonadal degeneration and a pronounced intestinal cell phenotype. A population of refractile, autofluorescent intestinal vesicles, which take up the lysosomal dye Neutral Red, increases dramatically in size, while the number of normal intestinal vesicles and the intestinal cytoplasmic volume decrease progressively. This is accompanied by an increase in the gut lumen size and a compromise in the intestine's ability to digest and absorb nutrients. CeTOR-deficient larvae exhibit no significant dauer characteristics, but share some features with starved L3 larvae. Notably, however, starved larvae do not have severe intestinal atrophy. Inactivation of C. elegans p70S6K or TAP42 homologs does not reproduce CeTOR deficiency phenotypes, nor does inactivation of C. elegans TIP41, a putative negative regulator of CeTOR function, rescue CeTOR deficiency. In contrast, inactivating the C. elegans eIF-4G homolog and eIF-2 subunits results in developmental arrest accompanied by the appearance of large, refractile intestinal vesicles and severe intestinal atrophy resembling that of CeTOR deficiency.. The developmental arrest and intestinal phenotypes of CeTOR deficiency are due to an inhibition of global mRNA translation. Thus, TOR is a major upstream regulator of overall mRNA translation in C. elegans, as in yeast.

Topics: Adaptation, Physiological; Alleles; Animals; Atrophy; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Eukaryotic Initiation Factor-2; Eukaryotic Initiation Factor-4G; Gene Expression Regulation, Developmental; Gene Targeting; Genes, Lethal; Genetic Complementation Test; Intestines; Larva; Phenotype; Phosphotransferases (Alcohol Group Acceptor); Protein Biosynthesis; Ribosomal Protein S6 Kinases, 70-kDa; RNA Interference; RNA, Messenger; Sequence Homology, Amino Acid; Sirolimus; Species Specificity; Starvation

Maturation arrest and interference with selection are two well-documented effects of cyclosporin-A (CsA) on the thymus. We recently hypothesized that these effects are related and owing to the reduced T-cell receptor (TCR)-CD3 complex-mediated signal transduction in thymocytes upon CsA treatment. In this hypothesis, the maturation arrest is the result of the additional depletion of thymocytes that normally survive by positive selection, whereas the impaired self-tolerance induction is caused by an increased survival of thymocytes that normally undergo negative selection. In this view, it is anticipated that CsA differentially affects thymocyte apoptosis during in vivo thymocyte maturation. Indeed, we report in this study a strong increase in apoptotic cells in the thymic cortex on in situ analysis. Simultaneously, the number of apoptotic cells had decreased at the cortico-medullary zone which is held to be the site for negative selection. Rapamycin (Rapa) also interferes with thymocyte maturation by inhibiting cytokine-driven proliferation. Hence, Rapa preferentially affects the early maturational stages of thymocyte development and is considered not to alter thymocyte selection and subsequent apoptotic events. Indeed, the number of apoptotic events appears not to be altered. However, possibly owing to the decrease in cortical macrophages, the apoptotic cells revealed an atypical enumeration around blood vessels. Taken together, our results favour the hypothesis that the dominant effect of CsA on the thymus is the reduction of the TCR-CD3 complex-mediated signal transduction in thymocytes upon interaction with stromal cells. Furthermore, the preferential localization of apoptotic cells next to blood vessels upon Rapa administration may indicate that endothelial cells are a back-up system for the removal of apoptotic cells.

Topics: Animals; Apoptosis; Atrophy; Cell Differentiation; Clonal Deletion; Cyclosporine; Endothelium, Vascular; Female; Immunosuppressive Agents; Models, Immunological; Rats; Rats, Inbred Lew; Receptor-CD3 Complex, Antigen, T-Cell; Self Tolerance; Signal Transduction; Sirolimus; Specific Pathogen-Free Organisms; T-Lymphocytes; Thymus Gland