Search

To ensure safety tolerance induction protocols are accompanied by conventional immunosuppressive drugs (IS). But IS such as calcineurin inhibitors (CNI), for example, cyclosporin A (CsA), can interfere with tolerance induction. We investigated the effect of an additional transient CsA treatment on anti-CD4mAb-induced tolerance induction upon rat kidney transplantation. Additional CsA treatment induced deteriorated graft function, resulting in chronic rejection characterized by glomerulosclerosis, interstitial fibrosis, tubular atrophy and vascular changes. Microarray analysis revealed enhanced intragraft expression of the B cell attracting chemokine CXCL13 early during CsA treatment. Increase in CXCL13 expression is accompanied by enhanced B cell infiltration with local and systemic IgG production and C3d deposition as early as 5 days upon CsA withdrawal. Adding different CNIs to cultures of primary mesangial cells isolated from glomeruli resulted in a concentration-dependent increase in CXCL13 transcription. CsA in synergy with TNF-α can enhance the B cell attracting and activating potential of mesangial cells. Transient B cell depletion or transfer of splenocytes from tolerant recipients 3 weeks after transplantation could rescue tolerance induction and did inhibit intragraft B cell accumulation, alloantibody production and ameliorate chronic rejection.

Recent data suggest that donor-specific memory T cells (T(mem)) are an independent risk factor for rejection and poor graft function in patients and a major challenge for immunosuppression minimizing strategies. Many tolerance induction protocols successfully proven in small animal models e.g. costimulatory blockade, T cell depletion failed in patients. Consequently, there is a need for more predictive transplant models to evaluate novel promising strategies, such as adoptive transfer of regulatory T cells (Treg). We established a clinically more relevant, life-supporting rat kidney transplant model using a high responder (DA to LEW) recipients that received donor-specific CD4(+)/ 8(+) GFP(+) T(mem) before transplantation to achieve similar pre-transplant frequencies of donor-specific T(mem) as seen in many patients. T cell depletion alone induced long-term graft survival in naïve recipients but could not prevent acute rejection in T(mem)(+) rats, like in patients. Only if T cell depletion was combined with permanent CNI-treatment, the intragraft inflammation, and acute/chronic allograft rejection could be controlled long-term. Remarkably, combining 10 days CNI treatment and adoptive transfer of Tregs (day 3) but not Treg alone also induced long-term graft survival and an intragraft tolerance profile (e.g. high TOAG-1) in T(mem)(+) rats. Our model allows evaluation of novel therapies under clinically relevant conditions.

The biologically active metabolite of vitamin D3, 1alpha,25-dihydroxyvitamin D3, has potent immunomodulatory activity; however, its clinical use is limited because of its hypercalcaemic activity in anti-inflammatory active doses. Here, we present ZK203278, a novel, structurally different vitamin D3 analogue with profound immunomodulatory activities. It potently inhibits lymphocyte proliferation in the mixed lymphocyte reaction, and release of cytokines that are central in inflammation, such as TNFalpha and IL-12 in the low nanomolar range. Similarly, expression of cell-surface molecules involved in cell adhesion and antigen presentation, e.g. to T cells, is down-regulated on human monocytes by low nanomolar concentrations of ZK203278. Potent anti-inflammatory activity has been demonstrated also in vivo in rodent disease models. ZK203278 inhibited allergic contact dermatitis in rodents after oral administration in doses approximately two orders of magnitude below the hypercalcaemic threshold dose. Moreover, ZK203278 significantly prolonged skin allograft survival in rats in well-tolerated doses. Altogether ZK203278, in contrast to 1alpha,25-dihydroxyvitamin D3, exerts considerable immunomodulatory activity at non-hypercalcaemic dosages and may have therapeutic potential for immune disorders or transplant rejection.

BACKGROUND Despite improved immunosuppression, intestinal transplantation is still complicated by severe rejection episodes. To further improve immunosuppressive concepts, we evaluated an anti-CD4 antibody and an anti-tumor necrosis factor (TNF)-alpha monoclonal antibody for their immunosuppressive efficacy in the standard rat model of intestinal transplantation. METHODS Intestinal transplantation was performed in the DA to Lewis combination, and recipients were treated perioperatively with either the anti-CD4 antibody RIB5/2 (day -1, 0, postoperative days 1, 2, 4, 7, 10, 14, 17, and 21), the anti-TNF antibody etanercept (60 min before reperfusion, postoperative days 3, 6, and 9) or a combination of both. Survival, histology and expression of immunologic mediator genes on days 3 and 4 after transplantation were investigated. RESULTS Treatment with anti-CD4 antibody alone (19.71+/-5.94) and the antibody combination (171.58+/-122.76) prolonged survival. The chemokine MIP-1alpha was significantly decreased in both anti-CD4 antibody treatment groups, possibly indicating an additional effect of the TNF-alpha blockade on the immune modulation by RIB5/2. CONCLUSIONS Our study demonstrated long-term graft survival in short-term treatment with a combination of an anti-CD4 antibody and a TNF-alpha antibody in more than 50% of the recipients of intestinal grafts. Such a combined approach could also be useful in clinical small bowel transplantation.

Zur gezielten Anwendung von Insektiziden gegen Borkenkafer an Ziergeholzen sind Pyrethroid-Praparate zugelassen, die in Streichapplikation aufgebracht werden sollen. Mit Hilfe eines im Handel angebotenen Kleingerates wurde vom Autor und von Garten- und Landschaftsbau- bzw. Baumschul-Unternehmen die Anwendung praktiziert und die gute Verwendbarkeit des technischen Hilfsmittels festgestellt. Auch die Anwendung eines zur Docht-Streichbehandlung zugelassenen Herbizides wurde erprobt. Okonomische und okologische Vorteile des Gerates im Sinne der guten fachlichen Praxis werden aufgezeigt.

This study examined whether a heart or kidney graft could provide protection for the more resistant skin graft. Buffalo rat recipients were given a single dose of RIB 5/2 (non-depleting anti-CD4 mAb) plus i.v. Lewis splenocytes 21 days before being given Lewis heart or kidney grafts. Lewis skin was grafted either simultaneously with, or after, long-term (>50 days) Lewis heart or kidney allograft acceptance. Immune responsiveness was analyzed by in vitro mixed lymphocyte culture (MLC), cytotoxic T lymphocytes (CTLs), and limiting dilution analysis (LDA). While i.v. alloantigen plus RIB 5/2 resulted in long-term acceptance of heart and kidney, survival of skin grafts alone was not prolonged. However, simultaneous transplantation with kidney, but not heart, resulted in long-term skin graft acceptance, while skin grafts subsequently grafted to recipients tolerant to kidney or heart were not accepted. In vitro analysis revealed a down-regulation of proliferation, cytotoxicity, and precursor T-helper cells (pThs)/precursor cytotoxic T lymphocytes (pCTLs) in Buffalo recipients accepting Lewis kidney and skin allografts. While RIB 5/2 plus Lewis splenocytes do not prolong the survival of skin grafts, Lewis skin grafted simultaneously with a kidney, but not heart, is accepted indefinitely and provides donor-specific protection for a subsequent skin graft.

The effectiveness and safety of fumaric acid esters (FAEs) for the treatment of psoriasis has been demonstrated. Their mode of action, however, is poorly understood. To determine the immunomodulatory potential of calcium methyl hydrogen fumarate (CaMHF) in transplant models, we tested the compound in rat transplantation models of acute rejection (AR) and chronic rejection (CR). Orthotopic kidney transplantation was combined with contralateral nephrectomy using the strain combinations WF to BDIX (AR) and F344 to LEW (CR). Recipients were treated orally prophylactically (day -28 to day +28, AR and CR model) or therapeutically (day +30 to day +60, CR model). CaMHF significantly prolonged the time to onset of AR in the WF/BDIX model. The half-lives of the grafts were 14 days in the CaMHF group, 7 days in the placebo-treated control group and 9 days in the untreated control group ( P0.01). Three of ten CaMHF-treated rats showed permanent graft acceptance (defined as survival for100 days) resulting in a mean survival time of42.3+/-41.0 ( P0.01) in comparison with28.3+/-38.3 days in the placebo-treated group and 9.4+/-2.6 days in the untreated control group. In the F344/LEW model of chronic graft injury, only prophylactic CaMHF treatment significantly inhibited the development of CR ( P=0.001; mean survival times28.4+/-2.0 weeks,23.9+/-6.0 weeks and21.1+/-5.4 weeks in the prophylactic CaMHF, therapeutic CaMHF, and placebo group, respectively). By 30 weeks six of ten prophylactically treated animals were still alive, but only three of nine and one of ten were alive in the therapeutically treated and placebo-treated groups, respectively ( P0.05). These findings indicate that CaMHF treatment effectively inhibits AR and CR, and demonstrates marked in vivo immunomodulatory efficacy. Thus, FAEs should be considered as drugs showing considerable immunosuppressive efficacy. This might have implications with regard to safety issues (e.g. immune monitoring) and novel potentially suitable indications (e.g. transplantation and other immune diseases).

Today, the major problem in organ transplantation is not acute graft rejection but chronic graft deterioration. In addition to alloantigen-specific events, alloantigen independent factors like donor age, previous diseases, consequences of brain death, and perioperative events of ischemia/reperfusion injury have a major impact on long-term graft function. The induction of the stress protein heme oxygenase-1 (HO-1) protects cells from injury and apoptosis. Here, we tested the protective effects of HO-1 induction in a clinically relevant kidney transplant model. Induction of HO-1 expression following cobalt-protoporphyrin (CoPP) treatment in organ donors prolonged graft survival and long-term function remarkably following extended periods of ischemia. Positive effects were observed with both optimal and marginal grafts from old donor animals. Structural changes characteristic for chronic rejection, as well as graft infiltration by monocytes/macrophages and CD8+ T cells, were substantially reduced following HO-1 induction. Up-regulation of HO-1 expression before organ transplantation was also associated with reduced levels for tumor necrosis factor (TNF)-alpha mRNA, increased levels for interferon (IFN)-gamma, and bcl-x, and insignificant differences for CD25, interleukin (IL)-2, IL-4, IL-6, and IL-10 mRNA levels. The significant improvement of long-term graft function following induction of HO-1 expression in donor organs suggests that this strategy may be a novel clinical treatment option with particular relevance for transplantation of marginal organs.

Objectives: heat shock protein (HSP70) has been studied in the ischaemic myocardium and proven to provide protection against ischaemia. However, HSP70 in ischaemic skeletal muscle in patients with peripheral arterial occlusive disease (PAOD) has not been reported. Methods: thirty-four patients with PAOD (Fontaine's criteria: stage II: 15; III: 9 and IV: 10, respectively) and ten non-PAOD controls were enrolled in the study. Calf muscle samples were taken. HSP70 was quantitated by SDS-PAGE using ultrasensitive silver staining with reference to a series of standard HSP70, and HSP70 mRNA was estimated using RT-PCR. Results: in comparison with the controls [median with range: 24.8 (14.1–35.6) ng in 2.5 μg total protein], HSP70 was increased significantly in PAOD [stage II: 93.1 (62.7–114.3); stage III: 110.1 (89.7–134.5) and stage IV: 77.4 (67.3–101.1)]. Similar results were obtained with HSP70 mRNA. Conclusions: HSP70 is increased in the ischaemic skeletal muscle in patients with PAOD, and HSP70 expression is different with regard to clinical stages, and the upregulation of HSP70 mRNA implies that the expression of HSP70 seems to be regulated at transcriptional level.

Leptin (L) is associated with body-weight-regulating and adipostatic functions. Its receptors also may be found centrally. Thyroid hormones regulate metabolic processes mainly by binding at peripheral receptors. Aim of this study was to show if there is a link between those central and peripheral regulation systems and to investigate the influence of different training intensities on L and the hypothalamic-thyroid-axis (HTA) in highly trained rowers. Six rowers (18.9 +/- 2.6 y; BMI 22.8 +/- 2.1 kg/m (2)) undertook high intensity resistance training (RT) for three weeks followed by three weeks of endurance training (ET). After each training cycle the subjects had one week for recovery (R1, R2). Blood samples were taken before and at the end of RT, R1, ET and R2. L, thyroid stimulating hormone (TSH), free T3 (fT3) and free T4 (fT4) were measured. After RT, a significant reduction in L, TSH and fT3 was found (p < 0.05). fT4 was unchanged. L remained decreased until the end of R1. After ET, a significant increase of TSH was found. L correlated to basal TSH levels (r = 0.49, p = 0.006) during R. BMI and body fat were unchanged throughout the study and were not correlated with hormonal levels. We speculate a high energy flux during intensified training (RT) caused the decrease of L and the HTA, independent of BMI or body fat. Thus, we conclude a depression of L and HTA is associated with training intensity.

The non-depleting anti-CD4 monoclonal antibody (mAb) RIB5/2 is a powerful inducer of tolerance to major histocompatibility complex (MHC)-incompatible allografts in rat recipients. The unresponsiveness induced is characterized by the persistence (over 300 days) of donor-reactive regulatory T cells within the graft. We applied differential-display reverse-transcription polymerase chain reaction (RT-PCR) to identify differences at the mRNA level between graft-infiltrating cells of anti-CD4 mAb-treated and non-treated control rats at day 5 after kidney transplantation. A 550-bp DNA fragment appearing only in anti-CD4 mAb-treated rats is identical with the anti-apoptotic protein Bag-1. A further investigation of Bag-1 expression during mixed lymphocyte reactions (MLR) revealed a three–four-fold up-regulation of Bag-1 mRNA expression in anti-CD4 mAb-treated allogeneic cultures. Bag-1 up-regulation is associated with higher protection against apoptosis of anti-CD4 mAb-treated cultures. Application of antisense oligonucleotides specific for Bag-1 leads to both a reduction in Bag-1 expression and sensibility against apoptosis. Thus, the expression of Bag-1 in anti-CD4 mAb-treated alloreactive T cells conferred resistance against apoptosis, which may contribute to the long-term survival of tolerance-mediating T cells in vivo.

Gene therapy is an interesting approach for the correction of defective genes, the treatment of cancer and the introduction of immunomodulatory genes. Various techniques for gene transfer into cells or tissues have been developed within the last decade; these can be divided generally into viral and nonviral gene transfer systems. Nonviral techniques include the liposome- or gene gun-mediated introduction of therapeutic genes; however, the efficiency of gene transfer by these applications is still very low. In contrast, viruses have optimised their strategies for efficient infection of virtually any cell type in a mammalian organism. The genetic modification of genomes from different virus families (Adenoviridae, Retroviridae, Herpesviridae) led to the development of gene therapy vectors with a similar capacity to infect cells or tissues as that of wild type viruses. In contrast to wild type viruses, gene therapy vectors are engineered to transfer therapeutic genes into the target cells or tissues. In addition, they have lost their capacity for replication in target cells, because of the removal of essential genes, which allows replication only in specialised packaging cell lines engineered for the production of recombinant viruses. Despite considerable progress over the past decade in the generation of gene transfer systems with reduced immunogenic properties, the remaining immunogenicity of many gene therapy vectors is still the major hurdle, preventing their frequent application in clinical trials. Recombinant adenoviruses have been shown to be promising vectors for gene therapy, since they are able to transduce both quiescent and proliferating cells very efficiently. However, a major disadvantage of adenoviral vectors lies in the activation of both the innate and adaptive parts of the recipient's immune system when applied in vivo. The inflammatory responses induced by adenovirus particles can be very strong and can be fatal in patients treated with these adenoviral constructs. Therefore, many experiments have been performed in the effort to prevent these inflammatory responses mediated by adenoviral particles. The depletion of cell populations responsible for these inflammatory responses as well as the application of immunosuppressive drugs have been investigated. Moreover, the generation of less immunogenic adenoviral vectors by further genetic modification within the adenoviral genome has led to vectors with reduced immunogenic properties. Both strategies to reduce inflammatory responses against adenoviral particles are discussed in this review.

CD4-targeted mAb therapy results in permanent acceptance of cardiac allografts in rat recipients, in conjunction with features of the infectious tolerance pathway. Although CD4+ T cells play a central role, the actual cellular and molecular tolerogenic mechanisms remain elusive. This study was designed to analyze in vitro alloimmune responses of T lymphocytes from CD4 mAb-treated engrafted hosts. Spleen, but not lymph node, cells lost proliferative response against donor alloantigen in MLR and suppressed test allograft rejection in adoptive transfer studies, suggesting compartmentalization of tolerogenic T cells in transplant recipients. A high dose of exogenous IL-2 restored the allogeneic response of tolerogenic T cells, indicating anergy as a putative mechanism. Vigorous proliferation of the tolerogenic T cells in in vivo MLR supports the existence of alloreactive lymphocytes in tolerogenic T cell repertoire and implies an active operational suppression mechanism. The tolerogenic splenocytes suppressed proliferation of naive splenocytes in vitro, consistent with their in vivo property of dominant immune regulation. Finally, CD45RC+ but not CD45RC− T cells from tolerant hosts were hyporesponsive to alloantigen and suppressed the proliferation of normal T cells in the coculture assay. Thus, nondeletional, anergy-like regulatory mechanisms may operate via CD4+CD45RC+ T cells in the infectious tolerance pathway in transplant recipients.

Autonomic dysreflexia presents a special situation in high-lesion spinal cord injury, however, intentionally or self-induced autonomic dysreflexia directly before or during competition to increase performance, so called 'boosting', is also being reported. In order to examine the influence of autonomic dysreflexia on plasma catecholamines, cardiocirculatory and metabolic parameters, 6 spinal cord injured wheelchair athletes with high-level lesions underwent wheelchair ergometry without (ST1) and with (ST2) autonomic dysreflexia. At the point of exhaustion significantly higher values for norepinephrine and epinephrine were observed in ST2 than in ST1. During autonomic dysreflexia a significantly higher peak performance (77.5 vs. 72.5 watt), higher peak heart rate (161 vs. 149 x min(-1)), and peak oxygen consumption (1.96 vs. 1.85 l x min(-1)), with comparable peak lactate (7.11 vs. 7.00 mmol x l(-1)) were reached on average. The blood pressure values in ST2 were partially hypertensive and higher than in ST1. In conclusion, autonomic dysreflexia, as a sympathetic spinal reflex, leads to a higher release of catecholamines during exercise. This results in higher peak performance, peak heart rate, peak oxygen consumption, and higher blood pressure values. The peak lactate, as an indicator of the anaerobic lactate metabolism, was unchanged. However, autonomic dysreflexia presents an unpredictable risk, caused predominantly by hypertensive blood pressure values, for high-lesion spinal cord injured persons at rest and more so during exercise; it is seen as a prohibited manipulation by the doping guidelines of the International Paralympic Committee.

E1-deleted adenoviral vectors are efficient vectors for somatic gene therapy. Recently, we have shown that intratracheal application of an adenoviral reporter construct leads to significant reporter gene expression in rat lungs within 24 h. In contrast, reporter gene expression in syngeneic rat heart transplants after adenovirus-mediated gene transfer was delayed. Since the adenovirus cannot replicate, down-regulation of the hCMV-IE promoter controlled reporter gene expression in initially infected cells by cytokines, which are released as a result of ischemia/reperfusion injury, might be involved. In order to investigate the role of proinflammatory cytokines, eg TNF-alpha in affecting hCMV-IE promoter-driven reporter gene expression, transient blockade of TNF-alpha was achieved by local co-application of an Ad-construct encoding for a soluble TNFRp55-Ig chimeric molecule in a syngeneic rat heart transplantation model. Co-application of the reporter construct together with the TNFRp55-Ig chimeric molecule significantly increased the early reporter gene expression after transplantation. Moreover, infiltration of inflammatory cells (T cells, macrophages, NK cells) and production of TNF-alpha in the transplant was markedly reduced. Our results indicate that: (1) proinflammatory cytokines are involved in down-regulation of reporter gene expression in ischemia/reperfusion injured tissues; and (2) inhibition of TNF-alpha might be a useful tool to increase early gene expression in gene therapy protocols, particularly in transplantation. Gene Therapy (2000) 7, 1238-1243.

We have previously reported that HSP70 in human skeletal muscle could be induced by training. However, whether HSP70 induction is dependent upon exercise volume or exercise intensity remains unknown. The aim of the present study was to investigate the relationship between HSP70 and training intensity in rowers. Fourteen well-trained male rowers were divided into two groups (group A, n = 6; group B, n = 8). Group A performed higher intensity exercise during 1st phase, whereas group B performed higher intensity exercise during 2nd training phase. Training volume in 2nd phase increased in both groups. Both training intensity and volume were reduced in 3rd phase. Muscle samples were taken from m. vastus lateralis by fine needle biopsy before training, at the end of the 1st, 2nd and 3rd training phases. HSP70 was quantitatively determined using SDS-PAGE with silver stain. In group A, HSP70 increased significantly from 38 +/- 12 etag before training to 59 +/- 16 etag at the end of the lst training phase (loaded total protein 2.5microg), and decreased afterwards. In group B, HSP70 increase (from 36 +/- 11 etag to 50 +/- 13 etag) in the 1st phase was significantly smaller, there was a further increase of HSP70 in the 2nd phase (60 +/- 14 etag). At the end of the training, HSP70 decreased in both groups. Thus, HSP70 response to training seems to be dependent upon exercise intensity.

Persons with spinal cord injury (SCI) are especially prone to atherogenesis. This is partly explained by an unfavourable lipoprotein profile in these individuals. The impairment of the sympathetic nervous system, and the fact that SCI subjects are subject to extreme physical inactivity, may have an influence on their lipid profile and lipoprotein(a) concentration. We made a detailed investigation of the lipid profile as well as serum levels of adrenaline and noradrenaline in 80 men with SCI ranging from tetraplegia to low paraplegia and in 16 control subjects. The lipid profile of tetraplegics was characterized by elevated very low-density lipoprotein cholesterol and triglyceride levels and reduced high-density lipoprotein levels. In contrast, paraplegics had significantly higher low-density lipoprotein and total cholesterol levels. Tetraplegics had lower and the low-lesion paraplegics had higher adrenaline and noradrenaline levels than the high-lesion paraplegics and the control subjects. High-lesion SCI subjects also showed an extreme reduction in VO2max. The lipoprotein profile was dependent on the injury level and serum catecholamine concentrations. The lower the noradrenaline values, the lower the high-density lipoprotein cholesterol. The low-density lipoprotein also correlated to catecholamines and particularly adrenaline values. Despite the correlation between lipoprotein(a) and adrenaline, no significant differences in lipoprotein(a) were found within SCI individuals as well as between SCI individuals and control subjects, indicating the predominantly genetic determination of lipoprotein(a) and thus the cardiovascular risk. Different serum catecholamine levels due to impairment of sympathetic nervous system and VO2max levels were observed in SCI subjects. This was associated with a higher lipid risk profile for cardiovascular diseases; however, the risk profile is dependent on the lesion level.

BACKGROUND: The aim of this study was to determine in an open trial if physical exercise in sleep apnea patients is safe and/or influences respiratory disturbance index (RDI). METHODS: After being treated 3 months or more with nasal CPAP for moderate to severe sleep apnea syndrome, eleven patients (1 female, 10 male, mean age 52.2 years) began a six-month period of supervised physical exercise twice a week, 2 hours each time. Before and after this period a Polysomnography without CPAP was recorded, along with a bicycle exercise test with lactate profile, echocardiography, body-weight, and body-height measurement. RESULTS: No adverse effects or cardiopulmonary problems were observed. There was no significant change in body weight with physical training; no significant difference in either min SaO2 nor mean SaO2; and no significant improvement in fitness. No adverse cardiopulmonary effects or problems were observed. There was a decrease of the RDI from 32.8 to 23.6 (p < 0.05), without a significant change in the REM-sleep portion of total sleep time (TST), NREM sleep, or TST. CONCLUSIONS: A prescription for mild to moderate exercise is safe in the management of sleep apnea, and, even in the absence of a fitness improvement, there occurred a decrease in RDI without a change in sleep architecture.

BACKGROUND Recently we have demonstrated that the nondepleting anti-CD4 monoclonal antibody (mAb) RIB5/2 induces long-term acceptance of kidney and heart allografts in all rat strain combinations tested. Cytokine gene expression studies by reverse transcriptase-polymerase chain reaction revealed a reversed intragraft interleukin (IL)-4/interferon-gamma ratio. Whether IL-4 mediated immune deviation contributes to transplantation tolerance is not clear so far. METHODS To learn more about the functional relevance of the relative IL-4 up-regulation, IL-4 was overexpressed in rat heart allografts by using ex vivo adenoviral gene transfer. The efficiency of gene transfer was analyzed by reporter gene assays as well by reverse transcriptase-polymerase chain reaction analysis of IL-4 mRNA expression. RESULTS The intragraft overexpression of IL-4 did not prolong the allograft survival compared with controls. Moreover, neutralization of IL-4 by OX81 mAb did not prevent tolerance induction by RIB5/2 treatment. CONCLUSIONS Anti-CD4 mAb-induced tolerance is associated with an intragraft type1/type2 shift, however, the up-regulation of IL-4 alone is neither sufficient nor essential to induce tolerance to cardiac allografts in a high-responder strain combination.

Background, CD4-targeted therapy or blocking of CD28-B7 T-cell costimulation may produce indefinite cardiac allograft survival in presensitized rats. This study analyzes the immune events associated with tolerance pathways after the blockade of activation signal 1 (CD4 monoclonal antibody [mAb]) or signal 2 (CTLA4Ig). Methods and Results. Lewis rats sensitized with Brown Norway skin grafts reject LBNF1 cardiac allografts in 300×10 6 of splenocytes. CD4 mAb therapy abolished the transcription of both T helper (Th)1 and Th2 cytokines compared with rejecting controls. In contrast, CTLA4Ig treatment resulted in a selective sparing of Th2-type cytokines. Surviving grafts in both groups were largely protected from signs of chronic rejection. Conclusions. CD4 mAb-induced blockage of activation signal 1 or CTLA4Ig-mediated blockage of co-stimulatory signal 2 may induce a true transplantation tolerance in sensitized rats, as documented by permanent graft acceptance and attenuation of chronic injury. The infectious pathway operates in a cell dose-dependent manner. Th2-type deviation in the graft itself is not required for tolerance maintenance, and it does not necessarily lead to chronic injury.

The purpose of this study was to investigate the influence of different injury levels in persons with spinal cord injury (SCI) on epinephrine (Epi) and norepinephrine (NE) at rest and during graded wheelchair exercise and the related changes in heart rate and O2 uptake (V˙o 2). Twenty tetraplegics (Tetra), 10 high-lesion paraplegics (HLPara), 20 paraplegics with SCI below T5 (MLPara), and 18 able-bodied, nonhandicapped persons (AB) were examined. Because of the higher level of interruption of the sympathetic pathways, Tetra persons showed lower Epi and NE at rest and only slight increases during exercise compared with all other groups; the Tetra subjects’ impaired cardiac sympathetic innervation caused restricted cardioacceleration and strongly reduced maximalV˙o 2. When compared with AB persons, HLPara had comparable NE but lower Epi levels as a result of partial innervation of the noradrenergic system and denervation of the adrenal medulla. MLPara subjects showed an augmented basal and exercise-induced upper spinal thoracic sympathetic activity compared with AB subjects. The increase in heart rate in relation toV˙o 2 was higher in HLPara because of a smaller stroke volume as a result of venous blood pooling. The different exercise response in persons with SCI is a result of the interruption of pathways in the spinal cord to the peripheral sympathetic nervous system in addition to the motor paralysis.

The purpose of this review was to give an answer to the question whether there are convincing data to support the hypothesis of an amino acid imbalance as one possible mechanism to explain overtraining syndrome. Animal studies point to an enhanced synthesis of the neurotransmitter 5-hydroxytryptamine through an amino acid imbalance at the blood-brain barrier with a preferable tryptophan uptake into the brain, resulting in premature fatigue. Human studies, however, show contradictory results, mainly because of nonstandardized methodology, so that a final conclusion cannot be made at present. BCAA supplementation in addition to standard carbohydrate ingestion during sustained exercise seems to be of no eminent advantage to delay fatigue. The overall results concerning the BCAA hypothesis to explain overtraining are inconclusive and require more controlled experimental research.

Purpose: The parasympathetic, Addison type, overtraining syndrome represents the dominant modern type of this syndrome. Beside additional mechanisms, an autonomic or neuroendocrine imbalance is hypothesized as underlying. Methods/Results: Several findings support this thesis. During heavy endurance training or overreaching periods, the majority of findings give evidence of a reduced adrenal responsiveness to ACTH. This is compensated by an increased pituitary ACTH release. In an early stage of the overtraining syndrome, despite increased pituitary ACTH release, the decreased adrenal responsiveness is no longer compensated. The cortisol response decreases. In an advanced stage of overtraining syndrome, the pituitary ACTH release also decreases. In this stage, there is additionally evidence for decreased intrinsic sympathetic activity and sensitivity of target organs to catecholamines. This is indicated by decreased catecholamine excretion during night rest, decreased β-adrenore-ceptor density, decreased β -adrenoreceptor-mediated responses, and increased resting plasma norepinephrine levels and responses to exercise. However, this complete pattern is only observed subsequent to high-volume endurance overtraining at high caloric demands. Conclusion: The described functional alterations of pituitary-adrenal axis and sympathetic system can explain persistent performance incompetence in affected athletes.

Background and Purpose —Epidemiological data link heavy snoring to an increased risk for stroke, an association often ascribed to hypertension and/or sleep apnea. The aim of this study was to determine whether obstructive hypopneas, central apneas, or obstructive apneas during sleep alter blood flow of the middle cerebral artery (MCA). Methods —Doppler sonography of the MCA was performed in conjunction with nightly polysomnography in 11 men and one woman. Results —A significant decline in blood flow occurred in 76% (169/223) of obstructive hypopneas and in 80% (98/123) of obstructive apneas, compared with only 14% (13/96) of central apneas ( P ≤.0001). While duration of events was not significantly different, MCA blood flow reductions were associated only with the duration of the obstructive hypopneas ( P ≤.01) and not with the duration of central ( P =.17) or obstructive ( P =.07) apneas. The magnitude of fall in arterial oxygen saturation from baseline correlated with a reduced blood flow with obstructive hypopneas but not with obstructive or central apneas. Conclusions —With obstructive hypopneas and obstructive apneas, MCA blood flow is more often decreased in comparison to central apneas. MCA blood flow reductions occur with longer obstructive hypopneas and with those hypopneas with greater falls in oxygen saturation. These observations indicate pathophysiology relevant to an increased risk for stroke in heavy snorers and patients with obstructive hypopneas and apneas.

Although CD4-targeted therapy abrogates acute rejection and may induce permanent graft acceptance in rodents, little is known about the mechanisms of long-term graft survival in these models. Recently, we have shown that treatment with a nondepleting anti-CD4 monoclonal antibody (mAb) (RIB-5/2) induces long-term survival of renal, heart, and skin allografts in strong major histocompatibility complex I/II incompatible rat strains. Here, we demonstrate that the development of major histocompatibility complex-specific and tissue-nonspecific tolerance rather than graft adaptation is responsible for long-term anti-CD4 mAb-induced transplant survival. Donor-specific but not third-party heart and pancreatic islet grafts were accepted permanently without adjunctive therapy in long-term kidney allograft recipients, and infusion of naive or alloimmune splenocytes failed to break the tolerant state. Interestingly, alloreactive T cells were not depleted in these long-term survivors, as ex vivo donor-specific mixed lymphocyte reaction was largely unaffected. The reverse transcriptase-polymerase chain reaction analyses of long-term renal allografts before and after donor-specific antigen challenge revealed no changes in CD3 mRNA level, but showed up-regulation of CD25, interleukin (IL) 2, interferon (IFN) gamma, IL-4, and IL-10 mRNA in the early phase, suggesting the presence of alloreactive T cells in tolerant rats. At later time points, the expression of IFN-gamma declined rapidly, whereas IL-4 persisted, resulting in a reversal of IFN-gamma/IL-4 ratio. Our data demonstrate the stability of anti-CD4 mAb-induced tolerance despite persistence of alloreactive T cells, suggesting the role of active tolerance-maintaining mechanisms. The T helper (Th) 1/Th2 shift may be involved in this regulatory process, as anti-CD4 mAb prevents acute graft-deteriorating rejection by effectively blocking Th1 responses, and well-functioning grafts may tolerize themselves by inducing regulatory cells.

Background . The monoclonal antirat CD4 antibody RIB 5/2 has been shown to modulate the CD4 glycoprotein without eliminating the affected T cells. We have shown that the administration of multiple doses of RIB 5/2 during the peritransplantation period prevents the rejection of rat kidney allografts. Methods . We compared the efficacy of a single intraperitoneal dose of 20 mg/kg RIB 5/2 plus donor antigen (25 × 10 6 spleen cells [SCs]) given by either an intrathymic or an intravenous route with the depleting anti-CD4 monoclonal antibody (mAb) OX-38 plus antigen 21 days before a major histocompatibility complex (MHC)-mismatched Lewis (RT1 1 ) to Buffalo (RT1 b ) rat cardiac or renal allograft. By delaying the transplantation for 21 days, recovery from the nonspecific effects of the antibody treatment allowed the demonstration of donor antigen-specific tolerance. Results . OX-38 mAb given with intrathymic SCs induced tolerance to heart but not kidney grafts, whereas OX-38 given with intravenous SCs failed to prolong survival of either heart or renal allografts. In contrast, RIB 5/2 mAb administration, when combined with alloantigen given by either intravenous or intrathymic routes, induced tolerance to heart allografts, whereas only alloantigen given by the intravenous route with RIB 5/2 resulted in tolerance to renal grafts. Conclusions . Concomitant administration of intravenous donor alloantigen and the modulation of CD4 + recipient cells by nondepleting RIB 5/2, rather than elimination of these CD4 + cells with depleting mAb OX-38, is a more potent method for the induction of allograft tolerance.

The use of the acetylene rebreathing method to estimate cardiac output (CO) during high-intensity exercise, which may be influenced by recirculation of acetylene, has not been validated. This study was designed to validate the acetylene rebreathing method to measure CO during high-intensity exercise using the direct Fick method. CO was measured at rest and during exercise at 25%, 50%, 75% and 90% of the nine subjects' maximum oxygen uptake (VO2max) by the direct Fick and acetylene rebreathing method. CO measured by the acetylene rebreathing method correlated with work rate (r = 0.90, P < 0.01) and with oxygen uptake (r = 0.94, P < 0.01). The correlation coefficient of CO between both methods was r = 0.91 (P < 0.01). There was no significant difference in CO measured by each method at rest as well as at each work rate. The difference in CO between each method was greater at lower CO than at higher CO. At 90% of VO2max, the CO measured by acetylene rebreathing was nearly identical to that measured by the Fick method. It can be concluded that acetylene rebreathing for measurement of CO is valid not only at rest but also during exercise, especially during high-intensity exercise.

The nondepleting monoclonal antirat CD4 antibody, RIB 5/2, has been shown to modulate, but not eliminate, the CD4 + T cells and to prolong survival of rat skin, renal, or cardiac allografts when serially administered after transplantation. In the present study, we compared the efficacy of recipient pretreatment with a single dose of nondepleting RIB 5/2 or depleting OX-38 anti-CD4 monoclonal antibody plus donor alloantigen given intravenously or intrathymically 21 days before transplantation on the survival of completely MHC-mismatched rat cardiac allografts. Intraperitoneal injection of a single dose (20 mg/kg) of RIB 5/2 resulted in a decrease in CD4 surface molecule expression on peripheral CD4 + T cells without cell elimination as shown by FACS analysis. The nonspecific effect of a single dose of RIB 5/2 mAb had resolved by 21 days after treatment as evidenced by the almost complete recovery of normal surface CD4 molecule expression. Cardiac allografts transplanted immediately or 21 days after a single dose of RIB 5/2 alone were uniformly acutely rejected. On the other hand, recipients treated with depleting anti-CD4 OX-38 (20 mg/kg) acutely rejected cardiac allografts transplanted 21 days later, but indefinitely accepted all grafts transplanted on the same day. In contrast, combined treatment with i.v. donor splenocytes (25x10 6 ) plus nondepleting RIB 5/2, but not with depleting anti-CD4 mAb, OX-38, resulted in survival for more than 100 days in 75% of recipients of donor specific, but not third party, cardiac allografts transplanted 21 days later. Irradiation (3000 rads) of the i.v. donor splenocytes combined with RIB 5/2 abrogated their tolerizing effect. When donor antigen was given intrathymically, both RIB 5/2 and OX-38 resulted in indefinite tolerance to cardiac allografts transplanted 21 days later. The failure of exogenous administration of high dose (180,000 IU/injection) rIL-2 for 10 days to reverse the unresponsiveness of i.v. SC plus RIB 5/2 pretreatment suggests that this tolerant state is not due to a deficiency of IL-2. In vitro studies showed marked inhibition of MLC responsiveness and cytolytic T cell activity in tolerant recipients that cannot be reversed by the addition of IL-2. Thus, pretransplant intravenous donor alloantigen combined with a dose of nondepleting anti-CD4 mAb, RIB 5/2, which alone has no significant effect, induced donor specific cardiac allograft tolerance.

E1-deleted adenoviral vectors are efficient vectors for somatic cell gene therapy, but transgene expression is limited in part by a cytotoxic T cell response directed against virally transduced cells. Moreover, the development of a neutralizing antibody response limits secondary gene transfer with these vectors. Therapy with a depleting anti-CD4 antibody permits prolonged transgene expression in the lung and liver of mice. Furthermore, transient depletion of CD4+ lymphocytes blocks neutralizing antibody production and therefore allows repeat administration and expression of E1-deleted recombinant adenovirus. In this study, we investigated the efficacy of a novel nondepleting anti-CD4 antibody (RIB 5/2) in a model of lung-directed gene therapy in outbred rats. Treatment with RIB 5/2 permitted prolonged reporter gene expression and reduced adenovirus-induced peribronchial and alveolar inflammation in the lung. Moreover administration of RIB 5/2 blocked the development of an anti-adenoviral neutralizing antibody response in the lung and permitted secondary administration and expression of a recombinant adenovirus. These data support the role of immunomodulation in prolonging in vivo transgene expression by recombinant adenovirus.

To test the overtraining-related "imbalanced amino acid hypothesis" (19), the influence of an unaccustomed average 103 % 4 wk -1 increase in training mileage (ITV) on performance and on serum levels of individual amino acids (AAs) was examined in distance runners and controlled by an unaccustomed average 152 % · 4 wk -1 increase in tempo-pace and interval runs (ITI). Two mmol · l -1 lactate performance (2 LP) increased, 4 LP stagnated and total running distance (TD) decreased in the incremental test during ITV - which may indicate an ITV-dependent overtraining - in contrast to an ITI-related increase in 2 LP, 4 LP and TD. The summed serum AAs decreased in ITV (2744 ± 534 vs 2933 ± 663 umol · l -1 ; p < 0.05) in contrast to an ITI-related increase (3541 ± 657 vs 3252 ± 885 umol · l -1 ; p < 0.05) with an average 29) % higher final summed AAs concentration during ITI (p < 0.05). During ITV 12 individual AAs decreased by 6 -17 %, 8 remained constant and 3 increased (Cys, Met, fTrp) by 6 - 19 %, as opposed to an ITI-related increase in 16 AA by 6 - 55 %. The observed IPV-related changes in serum AAs profile were smaller than after completing contests as a marathon, a 100 km-runoran ultra-triathlon. It may be concluded that the observed small changes in AAs profile or AAA/BCAA and AA/LNAA ratios only represent an epiphenomenon without recognizable influence on incremental test performance, since increases in fTrp/LNAA ratios (+ 28 % in ITV vs + 45 % in ITI) were found to be related both to performance impairment (ITV) and improvement (ITI).

The immunosuppressive effects of RIB-5/2, a nondepleting anti-rat CD4 monoclonal antibody (mAb), were analyzed in a well-defined model of accelerated cardiac allograft rejection. (LEW x BN)F1 hearts are rejected within 24 hours in LEW hosts presensitized with BN skin grafts at day -7. Treatment with RIB-5/2 mAb (3.5 mg/day i.v.) at days -7 and -1, prolonged cardiac allograft survival to the median of >62 days. The long-term recipients rejected acutely third-party (Wistar-Furth) test skin grafts, without an adverse effect on the survival of the original cardiac transplants. Lymphocytes harvested from mAb-treated hosts significantly decreased proliferative responses of donor cells in mixed leukocyte reaction. The cell activation and cytokine elaboration patterns were evaluated at the mRNA and protein levels by competitive template reverse transcriptase polymerase chain reaction and immunohistochemistry, respectively. Cardiac allografts in CD4 mAb-treated rats at 24 hours displayed reduced CD3, CD25, tumor necrosis factor-alpha, interleukin (IL)-1beta, IL-2, interferon (IFN)-gamma, and IL-10 mRNA levels as compared to those in rejecting grafts. Equal amounts of IL-4 mRNA were detected throughout in both animal groups; the expression of IL-10 mRNA increased progressively in the treated hosts. In contrast, IFN-gamma was consistently depressed after mAb therapy. The mRNA levels coding for CD3, CD25, tumor necrosis factor-alpha, IL-1-beta, and IL-2 genes were comparable in long-surviving and rejecting allografts. The staining for IL-2R, IL-2, and IFN-gamma was diminished, whereas the staining for IL-4 was either unaffected or enhanced in well-functioning grafts in RIB-5/2 mAb-treated hosts. The untreated recipients elicited strong circulating IgM allo-Ab response, which peaked around the time of cardiac rejection and then switched to IgG allo-Ab 4-7 days after heart transplantation. Treatment with RIB-5/2 mAb decreased IgM and prevented the switch into the IgG allo-Ab response. In conclusion, the ability of RIB-5/2 mAb treatment to combat accelerated rejection and to produce long-term graft acceptance is unprecedented in our experience in this model. These data provide new insights into the complexities of the cellular and humoral responsiveness, contributing to the the induction of donor-specific unresponsiveness in sensitized hosts. This study, along with our previous reports, indicate that an immune deviation in which intragraft Th1-type cytokines (primarily IFN-gamma) are diminished and Th2-type cytokines (IL-4 and IL-10) are maintained represents the common effector mechanism of CD4 mAb regimens in recipients of vascularized organ allografts.

Achieving tolerance or drug minimization after transplantation and thus preventing permanent immunosuppression with all the known severe side effects is the most important goal in transplantation medicine. In the last 20 years major progress has been made in understanding the tolerance underlying mechanisms and develop therapeutic strategies in small animal models. However, such knowledge could be rarely translated into the development of successful new therapeutic approaches in clinical transplantation. The success is limited by clinical challenges which are not present in our clean animal facilities such as 1) heterologous immunity - pathogen-specific memory T and B cells recognize alloantigens and boost the immune response towards the allograft, 2) pre-sensitization of recipients - presence of allo-specific memory T and B cells which are inert to most known therapeutic regimens. Thus we know now that we need more personalized treatment strategies according to the patient's immune reactivity. Such a strategy should combine three important aspects: i) an improved immune monitoring; ii) treatment which target memory cells and iii) strategies to reinforce regulation. We have established preclinical transplant models with preformed allo-reactive or pathogen-specific memory T cells in which we compare effectiveness of different treatment approaches combining depletional with regulatory approaches. Furthermore, we have performed a DNA microarray screen on samples of transplant patients and identified surface molecules specifically expressed by naive, central memory, effector memory or terminal differentiated effector memory (TEMRA) T cells. Using this approach we hope to develop antibodies, which specifically deplete effector memory and TEMRA cells but spare naive and central memory T cells. Such a treatment will be associated with less side effects e.g. infectious complications as compared to global depletion of T and B cells.

The new monoclonal antirat CD4 antibody RIB 5/2, which detects another epitope than those covered by W3/25 and MRC OX35, was tested for its immunosuppressive potency following skin allografting by using strain combinations with different genetic barriers in the MHC and genetic low- or high-responder background. High-dose and long-term therapy of the grafted rats led to a significant delay of the acute rejection (P < 0.01) in the strain combination Wistar Furth-to-BDX as well as in LEW1W-to-LEW1A. No significant prolongation of the mean allograft survival time was obtained for the high-responder rats (LEW1A-to-LEW1W). Cytofluorometric analysis revealed that RIB 5/2 exerts the immunosuppressive activity predominantly by modulation of the CD4 glycoprotein. Furthermore, the dependence of the humoral immune response against the mouse-globulins upon the administered protein quantity could be demonstrated.

The biologically active metabolite of vitamin D3, 1alpha,25-dihydroxyvitamin D3, has potent immunomodulatory activity; however, its clinical use is limited because of its hypercalcaemic activity in anti-inflammatory active doses. Here, we present ZK203278, a novel, structurally different vitamin D3 analogue with profound immunomodulatory activities. It potently inhibits lymphocyte proliferation in the mixed lymphocyte reaction, and release of cytokines that are central in inflammation, such as TNFalpha and IL-12 in the low nanomolar range. Similarly, expression of cell-surface molecules involved in cell adhesion and antigen presentation, e.g. to T cells, is down-regulated on human monocytes by low nanomolar concentrations of ZK203278. Potent anti-inflammatory activity has been demonstrated also in vivo in rodent disease models. ZK203278 inhibited allergic contact dermatitis in rodents after oral administration in doses approximately two orders of magnitude below the hypercalcaemic threshold dose. Moreover, ZK203278 significantly prolonged skin allograft survival in rats in well-tolerated doses. Altogether ZK203278, in contrast to 1alpha,25-dihydroxyvitamin D3, exerts considerable immunomodulatory activity at non-hypercalcaemic dosages and may have therapeutic potential for immune disorders or transplant rejection.

Ischemia/reperfusion injury leads to activation of graft endothelial cells (EC), boosting antigraft immunity and impeding tolerance induction. We hypothesized that the complement inhibitor and EC-protectant dextran sulfate (DXS, MW 5000) facilitates long-term graft survival induced by non-depleting anti-CD4 mAb (RIB 5/2). Hearts from DA donor rats were heterotopically transplanted into Lewis recipients treated with RIB 5/2 (20 mg/kg, days-1,0,1,2,3; i.p.) with or without DXS (grafts perfused with 25 mg, recipients treated i.v. with 25 mg/kg on days 1,3 and 12.5 mg/kg on days 5,7,9,11,13,15). Cold graft ischemia time was 20 min or 12 h. Median survival time (MST) was comparable between RIB 5/2 and RIB 5/2+DXS-treated recipients in the 20-min group with >175-day graft survival. In the 12-h group RIB 5/2 only led to chronic rejection (MST = 49.5 days) with elevated alloantibody response, whereas RIB 5/2+DXS induced long-term survival (MST >100 days, p < 0.05) with upregulation of genes related to transplantation tolerance. Analysis of the 12-h group treated with RIB 5/2+DXS at 1-day posttransplantation revealed reduced EC activation, complement deposition and inflammatory cell infiltration. In summary, DXS attenuates I/R-induced acute graft injury and facilitates long-term survival in this clinically relevant transplant model.

In order to improve endurance by exercise on a bicycle ergometer, both the interval method (I) (exertion and recovery phases alternate each minute) and the continuous method (constant exertion) can be employed. We examined the effects of both methods on the following parameters: heart rate, blood pressure, rate-pressure product, glucose, lactate, and catecholamine levels, and physical performance. Two groups of nine male patients were trained daily on a bicycle ergometer for 3.5 weeks. These patients had undergone coronary bypass surgery 24 and/or 26 days before the training started. The training heart rate was set at 86% of the individual maximum heart rate. In the last week of training, the exercise intensity in both patient groups, following either I or C regimen, was 20:121 W and 83 W respectively. The exercise training lasted 20 minutes with the following findings: (1) there were no significant differences in blood pressure, rate-pressure product, rates of glucose and catecholamines, and (2) there was a significantly higher rate of lactate in the second ten minutes of the I training. Before and after the training period, the patients were subjected to a multistage bicycle ergometer exercise test (sitting). The following results obtained after the training favor the I method: (1) patients' physical performance increased (+0.63 vs. +0.26 W/kg; p less than 0.001); (2) heart rate was lower at rest (-9 vs. -4 beats/min; p less than 0.04) and at 75 W (-12 vs. -2 beats/min; p less than 0.02); (3) rate-pressure product was lower at rest (-1675 vs. -291; p less than 0.04) and at 75 W (-2810 vs. -735; p less than 0.05); (4) rate of lactate was lower at 75 W (-0.83 vs. -0.33 mmol/l; p less than 0.04); (5) catecholamines were not lowered by I or C training, and no differences between the two groups could be observed. Exercise training according to the I method involves both the aerobic and anaerobic capacity of the organism, whereas exercise training according to the C method involves only oxidative capacity. After coronary bypass surgery, the I method is better suited to increase physical performance and is more effective in economizing the cardiac function.

The sleep of humans and animals is influenced by environmental and inner organic circumstances like the sunlight and circadian rythms of hormones. Sleep by itself can be divided in phases of REM (Rapid Eye Movement)- and Non-REM sleep. Both sleep phases influence physical parameters as heartrate, blood pressure, muscle tension and they seem to be both essential for the organism. The change of sleep phases is centrally controled in the brain. But at present no single population of neurons and no single transmitter can be made responsible for the control of REM- and Non-REM-sleep. Research of the last three decades however could show that neurons in the gigantic cellular field (FTG) in the pons mainly control REM-Sleep. This part of the brain has mainly cholinoceptive receptors and cholinergic transmitters like carbachol can enforce REM-sleep and anticholinergic substances like atropin can suppress REM-sleep.Unlike cholinergic substances the influence of adrenergic and monoaminercig substances in sleep control could not be proven on a neuronal level. However since several years some authors postulate the influence of aminergic substances in sleep control. One of their arguments is the strong relation of respiratory neurons and sleep neurons in the same area of the brain and for respiratory neurons the parasympathic as well as the sympathic influence is proven.

The evaluation of the clinical state of an athlete, e.g. of current trainability and of the diagnosis of overload and overtraining, is already one of the most complicated tasks in athletic medicine. Training is not only repetitive physical exercises, but also regular regeneration as an integral part of a successful training program. As already shown in several experimental studies, clinical, metabolic and hormonal findings, including the psychologically-related monitoring of stress and recovery, seem to reflect the clinical state of athletes.Such parameters can be used to monitor training and regeneration in athletes. However, there remains some uncertainty concerning reliability of such parameters for monitoring training and regeneration of elite athletes during periods preparatory to major events like World Championships.This review deals with some of the aspects of these important practical and scientifical questions based on the experience of several preparatory training camps in rowing. Rowing has to be seen as strenuous middle time endurance stress of 5.5 to 8.0 minutes’ duration, for which the athletes perform hughe training programs in which the monitoring of adaptation is essential to prevent long-term overtraining.

Definition, types, symptoms, findings, underlying mechanisms, and frequency of overtraining and overtraining syndrome have been described in an introductory article to the present volume. During the past 10 years, our increasing knowledge in this field has also been discussed in different original and review articles well as presented at the 1996 Memphis Overtraining and Overreaching in Sports Conference and summarized in a book project. Aim of this present overview, which was presented during the 1997 Reisensburg Castle workshop, is an additional up-dating of our knowledge considering mechanisms underlying overtraining-related performance incompetence in affected athletes with respect to further results obtained in this field during the past 2 years. Particular emphasis has been given to the time-course of regeneration subsequent to overtraining as far as it is known at present. From an operational standpoint, the thesis was followed that findings such as impairment of neuromuscular function depressed β-adrenergic receptor density related depressed lipolysis, glycogenolysis, glycolysis, and heart rate response as well as depressed intrinsic sympathetic activity depressed turnover in contractile proteins depressed adrenocortical and pituitary-hypothalamic responsiveness in an advanced stage or iron deficiency can explain performance incompetence in overtrained athletes, whereas appropriate regeneration should be indicated by their normalization.

From an operational standpoint, overtraining can be defined as stress > recovery (regeneration) imbalance, that is, too much stress combined with too little time for regeneration –In this context, stress summarizes all individual training, non-training, and competition-dependent stress factors,–Particularly, additional exogenous non-training stress factors, such as social, educational, occupational, economic, nutritional factors, travel, and endogenous factors (genetic predisposition) exacerbate the risk of a resulting overtraining syndrome in a completely individual manner .The term overtraining syndrome describes an impaired state of health which is caused by overtraining and characterized by particular findings.

The clinical success of new treatment strategies aiming on inducing permanent graft acceptance will rely on the ability to determine whether specific unresponsiveness to donor alloantigens has developed and for how long it is maintained. To identify markers for such posttransplant monitoring, genes differentially expressed by graft infiltrating leukocytes during tolerance induction or rejection after kidney transplantation in rats were compared. A subsequently performed full kinetic analysis in two different transplant models, kidney and heart, in two species, rat and mouse identified two markers (TOAG-1, alpha-1,2-mannosidase) with high specificity and reproducibility, which are highly expressed during induction and maintenance of acceptance, and downregulated during rejection. Expression level of these markers showed a strong positive correlation with graft function. In addition, expression of both genes was downregulated in the peripheral blood and the graft prior to rejection, suggesting that these markers may be useful for monitoring in clinical transplantation where peripheral blood is the most easily accessible patient sample. Interestingly, downregulation of TOAG-1 and alpha-1,2-mannosidase expression occurred in graft infiltrating cells and expression of both genes was also downregulated after T-cell activation in vitro.

In order to assess the role of heterologous immunity on tolerance induction (TI) by signal 1 modification, the influence of rat cytomegalovirus infection (RCMVI) on TI by a non-depleting monoclonal anti-CD4 mAb (monoclonal antibody) (RIB 5/2) in a rat kidney transplant (KTx) model was investigated. Orthotopic rat KTx (Dark Agouty (DA)-->Lewis (LEW)) was performed after TI with RIB 5/2 [10 mg/kg body weight (BW); day -1, 0, 1, 2, 3; i.p. (intraperitoneal route)]. RCMVI (5x10E5 Plaque forming units [PFU] i.p.) was simultaneously conducted to KTx, 50 days after KTx, and 14 days before and after KTx. RIB 5/2 induced robust allograft tolerance even across the high-responder strain barrier. RCMVI broke RIB 5/2-induced tolerance regardless of the time of RCMVI but did not induce acute graft failure during the 120 days follow-up. RCMVI induced a significant chronic deterioration of allograft function (p

Angiotensin-II (Ang-II) type 1 (AT(1)) receptor blockers may delay the progression of chronic allograft nephropathy (CAN). However, neither the optimal time for initiating AT(1) receptor blockade in order to delay CAN potentially nor the role of Ang-II type 2 (AT(2)) receptors under AT(1) receptor blockade is known. Both AT receptors can regulate p53 expression and apoptosis. We investigated what time of initiation with AT(1) blockers most effectively delayed CAN as well as the role of the AT(2) receptor, and how angiotensin receptor blockade affected apoptosis and its regulating factors in this context in a rat model. Kidneys of Fisher (F344) rats were transplanted into Lewis rats. Animals were treated with AT(1) (candesartan) and/or AT(2) (PD123319) receptor antagonists, a calcium channel blocker, or vehicle (treatment periods: day -7 before to week 24 after transplantation (long term), week 12 to week 24 (late), day -7 to day +5 (early)) and observed the animals for 24 weeks. Reduction of proteinuria, grade of CAN, and number of apoptotic cells was most pronounced in animals receiving long-term AT(1) receptor blockade. A combined AT(1)/AT(2) blocker treatment reduced CAN similarly to AT(1) blocker treatment alone. The number of apoptotic cells and the level of p53 mRNA were significantly lower in long-term AT(1) blocker-treated animals. In summary, AT(1) receptor blockade delayed the progression of CAN, particularly in animals treated long term. Reduction of apoptosis could be related to these beneficial effects. The AT(2) receptor does not appear to play an important role in CAN.

FTY720 is highly effective in various models of transplantation and autoimmunity. In order to find drugs that act synergistically with a tolerance-inducing nondepleting anti-CD4 mAb we studied this combination in a strong DA to LEW kidney transplantation model. Rats were treated with 0.3 mg/kg of FTY720 for 14 days and anti-CD4 mAb RIB5/2, alone or in combination. After kidney transplantation serum creatinine and blood lymphocyte counts were monitored. Immunohistology, ELISPOT and TaqMan trade mark -PCR analysis of biopsies were performed. Short-term application of RIB5/2 but not FTY720 induced long-term survival of kidney transplants. Moreover, the combination of FTY720 + RIB5/2 prevented tolerance induction. In the combination group serum creatinine levels increased 1 week after cessation of therapy and all rats died from uremia within 72 days. Intragraft immunohistology, ELISPOT and real-time RT-PCR analysis at day 21 demonstrated an enhanced T-cell infiltration and activation but a diminished up-regulation of protective genes in the grafts from recipients receiving the combination therapy. In contrast, delayed application of FTY720 to RIB5/2-treated rats did not interact with RIB5/2-induced tolerance. In summary, FTY720 is powerful in preventing intragraft infiltration by naive T cells but this might also affect the early development of graft-protecting regulatory T cells and tolerance induction.