Evaluation of clinical and safety outcomes associated with conversion from brand-name to generic tacrolimus in transplant recipients enrolled in an integrated health care system.

STUDY OBJECTIVE: To evaluate clinical and safety outcomes among transplant recipients whose tacrolimus was converted from the brand-name formulation to a generic formulation. DESIGN: Retrospective analysis. DATA SOURCE: Clinical databases and electronic records from a large, integrated health care system in California. PATIENTS: A total of 234 clinically stable, adult transplant recipients (renal, liver, and heart) whose tacrolimus was converted from the brand-name formulation to a generic formulation between October 1, 2010, and December 31, 2010, according to a physician-approved protocol. MEASUREMENTS AND MAIN RESULTS: For each patient, pre- and postconversion tacrolimus trough concentrations and serum creatinine concentrations were analyzed. Data were also collected on the percentage of patients who required dosage titration, drug cost savings, and rates of reversion to brand-name tacrolimus, biopsy-proved acute allograft rejections, and mortality. No significant differences were noted in mean ± SD pre- and postconversion tacrolimus trough levels (6.74 ± 1.61 vs 6.96 ± 2.31 ng/ml, p=0.137) or serum creatinine concentrations (1.33 ± 0.48 vs 1.36 ± 0.82 mg/dl, p=0.302). The mean ± SD percent change in tacrolimus trough concentration was 5.63 ± 32.95%. Thirty-six patients (15.4%) required dosage titration. Six patients (2.6%) reverted back to brand-name tacrolimus. No deaths or acute rejections occurred. Use of the generic product saved each patient an average of $45/month in drug acquisition cost and $26/prescription copayment. CONCLUSION: Clinical experience as well as research data show that use of generic tacrolimus results in trough concentrations that are comparable to the brand-name drug. Given the lack of adverse events reported and the cost savings recognized, conversion from brand-name tacrolimus to generic tacrolimus should be encouraged. Since dosage titration may be required, close therapeutic drug monitoring is recommended.

The smell of Tokishakuyaku-san (TJ-23) induces generation of regulatory T cells and prolongation of survival of fully allogeneic cardiac grafts in mice.

Oral administration of Tokishakuyaku-san (TJ-23), a Japanese herbal medicine, induces prolongation of cardiac allograft survival and generates regulatory cells in mice. Because herbal medicines usually have unique odor, and because smell is supposed to modulate the immune system, we examined whether the odor of TJ-23 induced prolonged allograft survival and regulatory cell generation. Naïve CBA mice (H2(k)) and olfactory-dysfunctional CBA mice after a stereotaxic operation underwent transplantation of C57BL/6 (B6, H2(b)) hearts, receiving fumigated water only or TJ-23 until rejection. Untreated or treated with water fumigation CBA mice rejected B6 cardiac grafts acutely (median survival times [MSTs], 7 and 8.5 days). When CBA mice were treated with fumigation of TJ-23, allograft survival was significantly prolonged (MST, 48 days). Olfactory-dysfunctional CBA mice treated with fumigation of TJ-23 rejected grafts acutely (MST, 7 days). Treatment with fumigation of TJ-23 also suppressed splenocytes proliferation and interferon-γ production. Secondary CBA recipients of whole splenocytes or CD4(+) cells from primary TJ-23-treated CBA recipients of B6 cardiac allografts at 30 days after grafting showed prolonged survival of B6 hearts (MST, >60 days). Flow cytometry studies showed increased CD4(+)CD25(+)Foxp3(+) regulatory cells in recipients given fumigation of TJ-23. In conclusion naïve but not olfactory-dysfunctional CBA mice treated with fumigation of TJ-23 displayed prolonged survival of fully allogeneic cardiac allografts and generation of regulatory cells.

Artemisiae capillaris herba induces prolonged survival of fully cardiac allografts and generates regulatory cells in mice.

Inchingorei-san (TJ-117), a 6-component herbal medicine, is used in Japan for the treatment of vomiting, urticaria, and liver and kidney disorders with few side effects. In this study we investigated the effect of TJ-117 on alloimmune responses in murine cardiac allograft transplantation. CBA (H2(k)) mice underwent transplantation of a C57BL/6 (B6, H2(b)) heart with oral administration of TJ-117 (or 1 component of TJ-117) from the day of transplantation for 7 days. CBA recipients given 1 g/kg/d of TJ-117 showed prolonged B6 allograft survival (median survival time [MST], 23 days). Naive CBA mice rejected B6 cardiac grafts acutely (MST, 7 days). Moreover, Artemisiae capillaris herba (ACH; 1g/kg/d) 1 component of TJ-117, significantly prolonged B6 allograft survival (MST, > 100 days). However, the other 5 components of TJ-117 were individually less effective than TJ-117 or ACH. ACH also suppressed splenocyte proliferation and interferon-γ production. Secondary CBA recipient showed prolonged survival of B6 hearts after treatments with whole splenocytes from primary ACH-treated CBA recipients carrying B6 cardiac allografts for 30 days (MST, >50 days). Flow cytometry studies showed increased CD4(+)CD25(+)Foxp3(+) regulatory cells in transplant recipients given ACH. In conclusion, ACH, 1 component of TJ-117, as well as TJ-117 induced hyporesponsiveness to fully allogeneic cardiac allografts with generation of CD4(+)CD25(+)Foxp3(+) regulatory cells.

Music exposure induced prolongation of cardiac allograft survival and generated regulatory CD4⁺ cells in mice.

In clinical practice, music has been used to decrease stress, heart rate, and blood pressure and to provide a distraction from disease symptoms. We investigated sound effects on alloimmune responses in murine heart transplantation. Naïve and eardrum-ruptured CBA/N (CBA, H2(K)) underwent transplantation of a C57BL/6 (B6, H2(b)) heart and were exposed to 1 of 3 types of music-opera (La Traviata), classical (Mozart), and New Age (Enya)-or 1 of 6 different single sound frequencies for 7 days. An adoptive transfer study was performed to determine whether regulatory cells were generated in allograft recipients. Cell-proliferation, cytokine, and flow cytometry assessments were also performed. CBA recipients of a B6 graft exposed to opera and classical music had significantly prolonged allograft survival (median survival times [MSTs], 26.5 and 20 days, respectively), whereas those exposed to 6 single sound frequencies and New Age did not (MSTs, 7, 8, 9, 8, 8, 8, and 11 days, respectively). Untreated and eardrum-ruptured CBA rejected B6 grafts acutely (MSTs, 7 and 8.5 days, respectively). Adoptive transfer of whole splenocytes, CD4(+) cells, and CD4(+)CD25(+) cells from opera-exposed primary recipients resulted in significantly prolonged allograft survival in naive secondary recipients (MSTs, 36, 68, and >50 days, respectively). Cell-proliferation, interleukin (IL)-2 and interferon-γ were suppressed in opera-exposed mice, whereas IL-4 and IL-10 from opera-exposed recipients were up-regulated. Flow cytometry studies showed an increased CD4(+)CD25(+)Foxp3(+) cell population in splenocytes from opera-exposed mice. In conclusion, exposure to some types of music may induce prolonged survival of fully allogeneic cardiac allografts and generate CD4(+)CD25(+)Foxp3(+) regulatory cells.

Auditory stimulation of opera music induced prolongation of murine cardiac allograft survival and maintained generation of regulatory CD4+CD25+ cells.

BACKGROUND: Interactions between the immune response and brain functions such as olfactory, auditory, and visual sensations are likely. This study investigated the effect of sounds on alloimmune responses in a murine model of cardiac allograft transplantation. METHODS: Naïve CBA mice (H2k) underwent transplantation of a C57BL/6 (B6, H2b) heart and were exposed to one of three types of music--opera (La Traviata), classical (Mozart), and New Age (Enya)--or one of six different single sound frequencies, for 7 days. Additionally, we prepared two groups of CBA recipients with tympanic membrane perforation exposed to opera for 7 days and CBA recipients exposed to opera for 7 days before transplantation (pre-treatment). An adoptive transfer study was performed to determine whether regulatory cells were generated in allograft recipients. Immunohistochemical, cell-proliferation, cytokine, and flow cytometry assessments were also performed. RESULTS: CBA recipients of a B6 cardiac graft that were exposed to opera music and Mozart had significantly prolonged allograft survival (median survival times [MSTs], 26.5 and 20 days, respectively), whereas those exposed to a single sound frequency (100, 500, 1000, 5000, 10,000, or 20,000 Hz) or Enya did not (MSTs, 7.5, 8, 9, 8, 7.5, 8.5 and 11 days, respectively). Untreated, CBA mice with tympanic membrane perforations and CBA recipients exposed to opera for 7 days before transplantation (pre-treatment) rejected B6 cardiac grafts acutely (MSTs, 7, 8 and 8 days, respectively). Adoptive transfer of whole splenocytes, CD4+ cells, or CD4+CD25+ cells from opera-exposed primary allograft recipients resulted in significantly prolonged allograft survival in naive secondary recipients (MSTs, 36, 68, and > 100 days, respectively). Proliferation of splenocytes, interleukin (IL)-2 and interferon (IFN)-γ production was suppressed in opera-exposed mice, and production of IL-4 and IL-10 from opera-exposed transplant recipients increased compared to that from splenocytes of untreated recipients. Flow cytometry studies showed an increased CD4+CD25+ Forkhead box P3 (Foxp3)+ cell population in splenocytes from those mice. CONCLUSION: Our findings indicate that exposure to opera music, such as La traviata, could affect such aspects of the peripheral immune response as generation of regulatory CD4+CD25+ cells and up-regulation of anti-inflammatory cytokines, resulting in prolonged allograft survival.

Induction of regulatory T cells and prolongation of survival of fully allogeneic cardiac grafts by administration of Tokishakuyaku-san in mice.

BACKGROUND: The Japanese herbal medicine Tokishakuyaku-san (TJ-23) has been used to treat neurodegenerative, immune, and respiratory tract diseases, as well as many gynecologic disorders, with few adverse effects. This study investigated the effect of TJ-23 on alloimmune responses in a murine model of cardiac allograft transplantation. METHODS: CBA mice underwent transplantation of a C57BL/6 heart and received oral administration of 2 g/kg per day of TJ-23 or 1 of 16 other commonly used Japanese herbal medicines from the day of transplantation until 7 days afterward. An adoptive transfer study was conducted to determine whether regulatory cells were generated. Histologic and cell proliferation studies, cytokine measurements, and flow cytometry analyses were also performed. RESULTS: Of the 17 herbal medicines studied, only TJ-23, given in a dose of 2 g/kg per day, induced significantly prolonged allograft survival (median survival time [MST], >100 days). TJ-23 also suppressed proliferation of splenocytes and production of interleukin-2, interleukin-6, and interferon-γ. Adoptive transfer of either whole splenocytes or CD4(+) or CD4(+) CD25(+) cells from TJ-23-treated allograft recipients resulted in indefinite survival of allografts in naive secondary recipients (MST >100 days). Flow cytometry studies showed that the CD4(+) CD25(+) forkhead/winged-helix (FOXP3)(+) regulatory cell population was increased in transplant recipients given TJ-23. CONCLUSION: TJ-23 induced hyporesponsiveness to fully allogeneic cardiac allografts and generated CD4(+) CD25(+) regulatory cells in our model.

Ursolic acid promotes robust tolerance to cardiac allografts in mice.

Nuclear factor (NF)-κB is an important molecule in T cell activation. Our previous work has found that T cell-restricted NF-κB super-repressor (IκBαΔN-Tg) mice, expressing an inhibitor of NF-κB restricted to the T cell compartment, can permanently accept fully allogeneic cardiac grafts and secondary donor skin grafts. In this study, we explore if transient NF-κB inhibition by a small molecular inhibitor could induce permanent graft survival. Ursolic acid, a small molecular compound, dose-dependently inhibited T cell receptor (TCR)-triggered NF-κB nuclear translocation and T cell activation in vitro. In vivo, ursolic acid monotherapy prolonged significantly the survival of cardiac allograft in mice. Assisted with donor-specific transfusion (DST) on day 0, ursolic acid promoted 84·6% of first cardiac grafts to survive for more than 150 days. While the mice with long-term surviving grafts (LTS) did not reject the second donor strain hearts for more than 100 days without any treatment, they all promptly rejected the third-party strain hearts within 14 days. Interestingly, this protocol did not result in an increased proportion of CD4(+) CD25(+) forkhead box P3(+) regulatory T cells in splenocytes. That adoptive transfer experiments also did not support regulation was the main mechanism in this model. Splenocytes from LTS showed reduced alloreactivity to donor antigen. However, depletion of CD4(+) CD25(+) regulatory T cells did not alter the donor-reactivity of LTS splenocytes. These data suggest that depletion of donor-reactive T cells may play an important role in this protocol.

Dietary vitamin K2 supplement improves bone status after lung and heart transplantation.

BACKGROUND: Osteoporosis is a problem after transplantation. Studies since the last year indicate that vitamin K plays a role in optimal bone health. The aim of this randomized, double blind, prospective longitudinal study was to investigate the effect of a dietary supplement with vitamin K2 (180 microg menakinon-7) on bone mass, the first year after lung and heart transplantation. METHODS: After preoperative baseline investigation of bone mass and bone-related biochemistry, 35 lung and 59 heart recipients were postoperatively randomized to vitamin K2 or placebo and reinvestigated the following year. RESULTS: In all recipients, 1 year after solid organ transplantation, the difference between vitamin K2 and placebo for the lumbar spine (L2-L4) bone mineral density (BMD) was 0.028 (SE 0.014) g/cm(2), P=0.055 and for L2 to L4 bone mineral content was 1.33 (SE 1.91) g/cm(2) (P=0.5). In lung recipients separately, the difference for bone mineral content was 3.39 g (SE 1.65), P=0.048 and in heart recipients 0.45 (SE 0.02) g, P=0.9 after controlling for baseline measures. In a forward stepwise linear regression analysis fitted to model differences in the L2 to L4 BMD, controlled for possible confounding variables (including use of bisphosphonate), and the only significant predictors were organ (B=-0.065 g/cm(2), P<0.001) and vitamin K2 (B=0.034 g/cm(2), P=0.019). Insufficient vitamin D status was common, and the parathyroid hormone was highest in the K2 group indicating a higher need for vitamin D. CONCLUSIONS: One year of vitamin K2 supplement suggest a favorable effect on lumbar spine BMD with different response in lung and heart recipients. Vitamin D status should receive more attention.

Protosappanin A induces immunosuppression of rats heart transplantation targeting T cells in grafts via NF-kappaB pathway.

Protosappanin A as one major and effective ingredient from Caesalpinia sappan L. exhibited antirejection activity obviously in heart-transplanted rat. The present study was designed to screen out the potential target genes of protosappanin A with microarray technology and reveal some molecular mechanism of immunosuppressive effect. Rats performed with ectopic peritoneal heart transplantation were randomized into three groups receiving different treatments for 7 days: protosappanin A group (25 mg kg(-1)), cyclosporine A group (10 mg kg(-1)), and control group. The differentially expressed genes responding to protosappanin A were analyzed with microarrays. Among common differentially expressed genes, the ones of interest were selected for further evaluation by real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), Western blot, immunochemistry, immunofluorescence, and ELISA. Among the 146 common differentially expressed genes, NF-kappaB and related genes like IkappaBa, IFN-r, and IP10 were selected for verification. The results of qRT-PCR, Western blot, immunochemistry, and ELISA showed that protosappanin A significantly reduced the expression of NF-kappaB, IFN-r, and IP10 (p < 0.05) and increased IkappaBa expression (p < 0.05) in graft. Moreover, the immunochemistry staining of NF-kappaB and IkappaBa was mainly observed in infiltrating mononuclear cells. Strikingly, immunofluorescent staining localized NF-kappaB to the TCR-positive T cells in graft. Furthermore, protosappanin A exhibited inhibitory effect on T cell proliferation in recipients after 7-day treatment. In conclusion, protosappanin A might act on T cells through inhibiting NF-kappaB activation and downstream gene expressions of IFN-r and IP10, meanwhile reducing T cell proliferation responding to alloantigen, so as to induce immunosuppressive effect. The results encourage a potential therapeutic evaluation of protosappanin A for clinical organ transplantation or other T cell-mediated immune disorders. Additionally, our study also verified the feasibility of microarray utilization in Chinese herb research to explore molecular mechanism and promote development of scientific theories.

Antirejection effect of Herba Asari extract in rats after cardiac allograft implantation.

OBJECTIVE: We studied the antirejection effect of asarinin, the extract of Herba Asari, in rats that underwent cardiac allograft implantation. METHODS: Rats received either olive oil (as a placebo control) or cyclosporine, with various amounts of asarinin or a combination of asarinin and cyclosporine daily through oral administration. 102 recipient SD rats were divided randomly into 7 groups: 1) group A, treated with 8 ml/kg olive oil per day; 2) group B, treated with 2.5 mg/kg cyclosporin A (CsA) each day; 3) group C, treated with 5 mg/kg cyclosporin A (CsA) each day; 4) group D1, treated with 12.5 mg/kg asarinin each day; 5) group D2, treated with 25 mg/kg asarinin each day; 6) group D3, treated with 50 mg/kg asarinin each day; 7) group E, treated with 12.5 mg/kg asarinin and 2.5 mg/kg CsA each day. Allograft survival was observed in each group of rats. On the seventh day posttransplantation; we analyzed weights, pathological lesions, and the levels of interleukin (IL)-2, interferon (IFN)-gamma, IL-4, and IL-10 in peripheral blood. RESULTS: The survival time of donor hearts in the asarinin groups was significantly prolonged (P < .01) when compared with the control group (7.8 +/- 0.7 days). The weights of rats increased in group C, group D2, group D3, and group E at 7 days after operation. Pathological lesions were significantly less severe. The levels of IL-2 and IFN-gamma decreased remarkably (P < .01), while those of IL-4 and IL-10 were not affected (P > .05) in the asarinin groups (D2 and D3). CONCLUSIONS: Asarinin decreased peripheral blood concentrations of IL-2 and IFN-gamma with prolongation of allograft heart survival.

Synergistic effects of Isatis tinctoria L. and tacrolimus in the prevention of acute heart rejection in mice.

Although immunosuppressive treatments are available for acute cardiac rejection no viable treatment exists for long-term cardiac graft failure. Moreover, the extended use of calcineurin inhibitor immunosuppressants, the mainstay of current treatment for cardiac transplantation, leads to significant side effects such as nephrotoxicity and an increased risk of cardiac disease. Because some agents used in Traditional Chinese Medicine (TCM) have strong immunosuppressive effects coupled with low toxicity, we investigated the effect of Compound K (K), the synthesized analogue of highly unsaturated fatty acids from Isatis tinctoria L., either as a single treatment or combined with tacrolimus (FK-506) on acute cardiac allograft rejection. We compared the ability of K alone, or in combination with FK-506, to inhibit acute heart transplant rejection both in vitro and in vivo. We found that the inhibition of lymphocyte proliferation was positively correlated with K concentration. K significantly reduced IL-2 and IFN-gamma expression levels and significantly inhibited lymphocyte proliferation in both a lymphocyte transformation test and a mixed lymphocyte reaction (MLR). We also found that the inhibitory effect of a combination of K and a sub-therapeutic dose of FK-506 (SubFK-506) was stronger than that of full-dose FK-506 alone. Oral administration of K reduced acute cardiac allograft rejection in mice and had no apparent toxicity. In vivo, the immunosuppressive effect of K combined with a half-dose of FK-506 was equivalent to that of a full-dose of FK-506 alone. K combined with a half-dose of FK-506 reduced the expression levels of IL-2 and IFN-gamma (both within the graft and in the recipients' serum) more effectively than a full-dose of FK-506. These results show that K has significant immunosuppressive effects both in vitro and in vivo. When used as a combination therapy with FK-506 we see a powerful inhibition of rejection with no obvious toxic side effects. The mechanism of action is postulated to involve the inhibition of IL-2 and IFN-gamma expressions by lymphocytes, rather than the activation of Tr1 cells via the production of IL-10.

Prolonged survival of fully mismatched cardiac allografts and generation of regulatory cells by Sairei-to, a Japanese herbal medicine.

Sairei-to (TJ114), a 12-component Japanese herbal medicine, is used to treat immune-related diseases. We investigated the effects of oral administration of TJ114 in a murine model of cardiac transplantation with fully mismatched allografts. Untreated CBA mice rejected C57BL/6 hearts acutely (median survival time [MST], 7 days), whereas survival of allografts from mice given TJ114 was significantly prolonged (MST >100 days). Secondary CBA recipients of C57BL/6 hearts also had prolonged allograft survival (MST >100 days) after adoptive transfer of whole or CD4 splenocytes from primary CBA allograft recipients given TJ114. None of the individual components of TJ114 prolonged allograft survival, suggesting that its effects require administration of the combination agent. In mixed leukocyte cultures, proliferation of splenocytes from TJ114-treated CBA recipients was markedly suppressed compared with that of splenocytes from untreated mice, and interferon-gamma production was significantly reduced. Thus, in our model, TJ114 treatment induced hyporesponsiveness to cardiac allografts and generated CD4 regulatory cells.

Green tea extract prolongs allograft survival as an adjunctive therapy along with low dose cyclosporine A.

The current immunosuppressive drugs are successful in prolonging allograft survival but fail to achieve transplantation tolerance or prevent chronic rejection. Consequently, there is ongoing research to develop novel combinatorial therapies that are more efficacious in prolonging allograft survival as well as induce tolerance toward the transplanted organ. The present study aims to study the efficacy of green tea extract (GTE) in combination with low dose cyclosporine A (CyA) in prolonging allograft survival in mice. Numerous studies have reported the anti-inflammatory and immunomodulatory properties of GTE and its various catechin components. GTE is also known to attenuate CyA induced nephrotoxicity. Therefore, we hypothesized that GTE alone or in combination with CyA will prolong graft survival. Our study demonstrates that GTE in combination with low dose CyA significantly prolongs graft survival as well as increase the production of immunosuppressive cytokine, IL-10. GTE also decreases CyA induced high TGF-beta production, which is incriminated in CyA induced nephrotoxicity. We also observed that GTE inhibits both nonspecific and antigen-specific proliferation of T cells in vitro. These results indicate the potential of GTE as an adjunctive therapy in combination with CyA to prolong allograft survival and to reduce CyA induced nephrotoxicity.

Protosappanin a, an immunosuppressive constituent from a Chinese herb, prolongs graft survival and attenuates acute rejection in rat heart allografts.

OBJECTIVE: Caesalpinia sappan L., which has been used in oriental medicine as an analgesic and antiinflammatory agent, has shown immunosuppressive activity on acute rejection on rat heart allografts. The present study was designed to investigate the potency of protosappanin A, one major ingredient of C. sappan L., in rat heart transplantation. MATERIALS AND METHODS: Protosappanin A isolated from an ethanol extract of C. sappan L. was identified by wave spectrum. All groups consisted of Wistar donors into Sprague-Dawley (SD) recipients, including large (25 mg/kg) or small (5 mg/kg) doses of protosappanin A plus cyclosporine (10 mg/kg). A control group was used. Animals were given the drugs on postoperative day 2. We observed graft survival and pathologic conditions of the hearts. Blood samples were obtained on postoperative day 7 to measure the CD4+/CD8+ T-cell ratio. Graft perforin and granzyme B mRNA expression levels were examined with reverse transcription-polymerase chain reaction (RT-PCR) methods. RESULTS: Protosappanin A or cyclosporine significantly prolonged heart allograft survival (P < .01), alleviated myocardial pathologic damages (P < .01), decreased the CD4+/CD8+ ratio (P < .05), and inhibited perforin and granzyme B mRNA expressions in the graft (P < .05). CONCLUSIONS: These findings demonstrated that protosappanin A prolonged heart allograft survival by significantly attenuating acute rejection.

[Effect of ethyl acetate extract of sappan wood on expression of myocardial GrB mRNA in rat model of allogeneic ectopic cardiac transplantation].

OBJECTIVE: To explore the action mechanism of the immune active components of sappon wood (SWE) for antagonizing reject reaction by observing the influence of its ethyl acetate extract on mRNA expression of myocardial GrB in rat model of allogenic ectopic cardiac transplantation. METHODS: Animal model of abdominal cardiac ectopic transplantation was established taking Wistar rat as the donor and SD rat as the receptor. The successfully modeled rats were randomly divided into the model group, the SWE group and the CsA group. GrB mRNA expression was detected by RT-PCR method and myocardial pathomorphologic picture was observed in routine. RESULTS: The pathologic changes in the SWE group (23 scores) and the CsA group (14 scores) were milder than in the model group (31 scores), the former two could markedly alleviate the myocardial pathologic injury (P<0.05, P<0.01). The GrB mRNA expression in the model group was 1.3000 +/- 0.1207, the SWE group 0. 7070 +/- 0.1215, and the CsA group 0.6700 +/- 0.0997, respectively; compared with the model group, the latter two could obviously down-regulate the expression of GrB mRNA (P<0.01) and no significant difference was found between the latter two groups (P>0.05). CONCLUSION: SWE could alleviate the pathologic change, down-regulate the mRNA expression of myocardial GrB in allogenic ectopic transplanted myocardium of rats, it is possibly one of the factors for its antagonizing effect against reject reaction.

Mechanisms underlying blockade of allograft acceptance by TLR ligands.

Immune activation via TLRs is known to prevent transplantation tolerance in multiple animal models. To investigate the mechanisms underlying this barrier to tolerance induction, we used complementary murine models of skin and cardiac transplantation in which prolonged allograft acceptance is either spontaneous or pharmacologically induced with anti-CD154 mAb and rapamycin. In each model, we found that prolonged allograft survival requires the presence of natural CD4(+)Foxp3(+) T regulatory cells (Tregs), and that the TLR9 ligand CpG prevents graft acceptance both by interfering with natural Treg function and by promoting the differentiation of Th1 effector T cells in vivo. We further demonstrate that although Th17 cells differentiate from naive alloreactive T cells, these cells do not arise from natural Tregs in either CpG-treated or untreated graft recipients. Finally, we show that CpG impairs natural Treg suppressor capability and prevents Treg-dependent allograft acceptance in an IL-6-independent fashion. Our data therefore suggest that TLR signals do not prevent prolonged graft acceptance by directing natural Tregs into the Th17 lineage or by using other IL-6-dependent mechanisms. Instead, graft destruction results from the ability of CpG to drive Th1 differentiation and interfere with immunoregulation established by alloreactive natural CD4(+)Foxp3(+) Tregs.

C. sinensis ablates allograft vasculopathy when used as an adjuvant therapy with cyclosporin A.

Immunosuppressive treatments are available to suppress acute cardiac rejection; however, no viable treatment exists for long-term cardiac graft failure. Moreover, extended use of calcineurin inhibitor immunosuppressants, the mainstay of the current therapeutic for cardiac transplantation, leads to significant associated pathologies such as nephrotoxicity and increased risk of cardiac disease. For the last ten years alternatives to calcineurin inhibitors, or adjuvant therapies designed to complement their activities, have been explored. In tandem with this development, there has been considerable interest in Traditional Chinese Medicines (TCM) as sources for novel therapeutics. Our study examines the ability of the TCM Cordyceps sinensis to reduce acute and chronic rejection associated with cardiac transplantation. The objectives of this study were to first determine if oral delivery of the extract could reduce acute rejection in a rat heterotopic heart model of transplantation. The second objective was to determine, in vitro, if a sterile, aqueous extract of C. sinensis could decrease CD8+ T cell activity. The third objective was to determine if oral delivery of the extract could ablate allograft vasculopathy in a mouse abdominal aortic transplant model. We found that oral delivery of the extract demonstrated a reduction in acute rejection when used in conjunction with a sub-therapeutic dose of Cyclosporine. Further, we found, using a mixed lymphocyte reaction, that the extract was able to significantly reduce CD8+ T cell activity. Finally, we demonstrate that oral delivery of the extract, used with a therapeutic dose of Cyclosporine to suppress acute rejection, ablates allograft vasculopathy.

Novel method for selecting immunosuppressive histone deacetylase (HDAC) inhibitors with minimal thrombocytopenia.

Histone deacetylase (HDAC) inhibitors repress interleukin-2 (IL-2) gene expression in T cells and possess immunosuppressive activity in vivo. In addition to its immunosuppressive activity, HDAC inhibitors block GATA binding protein-1 (GATA-1) gene expression in megakaryocytes and elicit thrombocytopenia. In this report we state that for a given immunosuppressive dose of HDAC inhibitor, the ratio of GATA-1 reporter gene activity relative to IL-2 reporter gene assay (G/I ratio of measured IC(50)) can be predictive of a HDAC inhibitor's thrombocytopenic effect. This study utilized nine HDAC inhibitors at a minimal effective dose in a rat heterotopic cardiac transplantation model and the resultant G/I ratios and platelet depletion rates were highly correlated (r=0.933). These results indicate that calculation of G/I ratio can be a novel method for selecting immunosuppressive HDAC inhibitor having minimal thrombocytopenic effect which will benefit the search for new immunosuppressants of greater safety and efficacy.

A comparison of the effects of fish oil and flaxseed oil on cardiac allograft chronic rejection in rats.

Both fish and flaxseed oils are major sources of different n-3 fatty acids. Beneficial effects of fish oil on posttransplantation complications have been reported. The current study aimed to compare the effects of flaxseed and fish oils in a rat cardiac allograft model. Male Fischer and Lewis rats were used as donors and recipients, respectively, to generate a heterotopic cardiac allograft model. Animals were randomly assigned into three groups and fed a diet supplemented with 1) 5% (wt/wt) safflower oil (control, n = 7), 2) 5% (wt/wt) flaxseed oil (n = 8), or 3) 2% (wt/wt) fish oil (n = 7), and an intraperitoneal injection of cyclosporine A (CsA; 1.5 mg.kg(-1).day(-1)) over 12 wk. Body weight, blood pressure, plasma levels of lipids, CsA, select cytokines, as well as graft function and chronic rejection features were assessed. Body weight and blood CsA levels were similar among the groups. Relative to controls, both treated groups had lower systolic and diastolic blood pressure and plasma levels of macrophage chemotactic protein-1. Treatment with fish oil significantly (P < 0.05) lowered plasma levels of triglycerides, total cholesterol, and LDL-cholesterol. HDL-cholesterol concentrations were significantly higher (P < 0.05) in the flaxseed oil-treated group compared with the other two groups. Both flaxseed oil and fish oil may provide similar biochemical, hemodynamic, and inflammatory benefits after heart transplantation; however, neither of the oils was able to statistically significantly impact chronic rejection or histological evidence of apparent cyclosporine-induced nephrotoxicity in this model.

Cardiac allograft vasculopathy after cardiac transplantation and hormone therapy: positive effects?

BACKGROUND: There is a great deal of controversy surrounding the issue of hormone replacement therapy after transplantation. The question whether or not this therapy has effects in cardiac allograft vasculopathy (CAV), the Achilles heel of cardiac transplantation or other unique aspects of allograft function is still unknown. METHODS: We investigated the long-term effect of 17beta-estradiol as well as phytoestrogen Coumestrol, a synthetically produced phytoestrogen, on the development of CAV and the degree of fibrosis in an ovariectomized female heterotopic chronic allograft model (LEW-F344). RESULTS: We found that, 150 days after transplantation, no significant effect of estrogen application on intimal thickening of coronary arteries was observed. 17beta-estradiol and phytoestrogen Coumestrol did significantly reduce the perivascular immune reaction. However, the immune effect had no consequence on the intensity of CAV. Although neither 17beta-estradiol nor phytoestrogen Coumestrol revealed a positive effect on CAV, the group of animals treated with 17beta-estradiol showed the highest decline in heart function and the most distinct fibrosis. CONCLUSIONS: 17beta-estradiol does not affect CAV positively, but worsens cardiac allograft function and leads to increased fibrosis. This is the first study showing a negative effect of 17-beta-estradiol after heart transplantation in the long term.