Incidence, diagnosis and treatment of popliteal artery entrapment syndrome in current vascular practice in Japan.

Few data regarding popliteal artery entrapment syndrome (PAES) is available in Japan. In this study, we investigated incidence, diagnosis and treatment of PAES in current vascular practice. A retrospective analysis of all patients with PAES visiting 31 participating institutes between 2003 and 2015 was conducted. Thirty-five limbs (28 patients) were identified during the 13-year study period, and the incidence of PAES was 0.12% of all peripheral artery disease cases revascularized. Mean age was 32.0 ± 16.9 years old, and 60 and more years old was 10.7%. Also, 92.9% were male and 39.3% were athletes. Most frequent initial symptoms were intermittent claudication in 23 limbs (65.7%); 4 limbs (11.4%) had chronic limb-threatening ischemia. CT scan was most frequently (94.3%) used for the diagnostic imaging followed by MRI (45.7%) and duplex ultrasound (45.7%). Stress test such as dorsal flexion during duplex ultrasound was used only in 28.6%. Thirty-two limbs (91.4%) received surgical treatment, including 23 arterial reconstructions (71.9%); there were no major perioperative complications. All patients achieved improvement of their symptoms, and the average ankle brachial index increased from 0.69 ± 0.22 to 1.00 ± 0.14 post-surgery. The average postoperative follow-up period was 26.0 months with only one reintervention during the follow-up. In conclusion, PAES was a rare condition and traditional surgical treatment was solid. However, given a broad spectrum of clinical feature of PAES and less usage of diagnostic duplex ultrasound with stress test, there might be a miss- or delayed diagnosis of PAES even in the current vascular practice.

Rapid detection of quality of Japanese fermented soy sauce using near-infrared spectroscopy.

This study investigated the feasibility of rapidly evaluating the final quality of Japanese fermented soy sauce (shoyu) using NIR spectroscopy and partial least-squares (PLS) regression. In total, 110 shoyu samples that had been entered in the annual soy sauce competition from 2016 to 2018 were collected and analyzed. The transmittance spectra (400-1800 nm) and the transflectance spectra (680-2500 nm) of these samples were acquired and processed by different pre-treatments. PLS regression was applied to the raw and processed spectra to construct models based on a calibration set (76 shoyu samples from 2016 and 2017) and to evaluate these models using a validation set (34 shoyu samples from 2018), according to their values for bias and root mean square error of prediction (RMSEP). The results showed that the models constructed using the full spectra of transflectance performed better than those using transmittance spectra. Comparing the influence of different regions in the transflectance spectra enabled the accuracy of the models to be improved. The model constructed from transflectance spectra from the 1800 to 2500 nm region using pre-treatment of second derivative was superior to the other models, with a bias value of -2 and the lowest RMSEP value of 13 in the validation set. To further narrow the wavelength range, the models constructed using the spectral region from 2050 to 2400 nm also showed a better performance for predicting the sensory quality of soy sauce products. This study has demonstrated that the NIR spectroscopy technique could be used as an alternative routine quality control procedure, which can rapidly and economically classify the quality of soy sauce products.

Frying stability of rapeseed Kizakinonatane (Brassica napus) oil in comparison with canola oil.

This study was carried out to investigate the frying performance of Kizakinonatane (Brassica napus) oil during deep-fat frying of frozen French fries with/without replenishment. Commercial regular canola oil was used for comparison. The frying oils were used during intermittent frying of frozen French fries at 180, 200, and 220 ℃ for 7 h daily over four consecutive days. The Kizakinonatane oil exhibited lower levels of total polar compounds, carbonyl value, and viscosity as well as comparable color (optical density) values to that of the canola oil. The monounsaturated fatty acid/polyunsaturated fatty acid ratios were lower than that of canola oil, whereas the polyunsaturated fatty acid/saturated fatty acid ratios are higher than that of canola oil after heating. Results showed that fresh Kizakinonatane oil contains higher levels of acid value, viscosity, optical density values, tocopherols, and total phenolics contents than that of canola oil. Replenishment with fresh oil had significant effects on all chemical and physical parameters, except the acid value of the Kizakinonatane oil during frying processes. Based on the results, the Kizakinonatane oil is inherently suitable for preparing deep-fried foods at high temperatures.

Both rejection and tolerance of allografts can occur in the absence of secondary lymphoid tissues.

In this study, we showed that aly/aly mice, which are devoid of lymph nodes and Peyer's patches, acutely rejected fully allogeneic skin and heart grafts. They mounted potent inflammatory direct alloresponses but failed to develop indirect alloreactivity after transplantation. Remarkably, skin allografts also were rejected acutely by splenectomized aly/aly (aly/aly-spl(-)) mice devoid of all secondary lymphoid organs. In these recipients, the rejection was mediated by alloreactive CD8(+) T cells presumably primed in the bone marrow. In contrast, cardiac transplants were not rejected by aly/aly-spl(-) mice. Actually, aly/aly-spl(-) mice that spontaneously accepted a heart allotransplant and displayed donor-specific tolerance also accepted skin grafts from the same, but not a third-party, donor via a mechanism involving CD4(+) regulatory T cells producing IL-10 cytokine. Therefore, direct priming of alloreactive T cells, as well as rejection and regulatory tolerance of allogeneic transplants, can occur in recipient mice lacking secondary lymphoid organs.

Primary vascularization of allografts governs their immunogenicity and susceptibility to tolerogenesis.

We investigated the influence of allograft primary vascularization on alloimmunity, rejection, and tolerance in mice. First, we showed that fully allogeneic primarily vascularized and conventional skin transplants were rejected at the same pace. Remarkably, however, short-term treatment of mice with anti-CD40L Abs achieved long-term survival of vascularized skin and cardiac transplants but not conventional skin grafts. Nonvascularized skin transplants triggered vigorous direct and indirect proinflammatory type 1 T cell responses (IL-2 and IFN-γ), whereas primarily vascularized skin allografts failed to trigger a significant indirect alloresponse. A similar lack of indirect alloreactivity was also observed after placement of different vascularized organ transplants, including hearts and kidneys, whereas hearts placed under the skin (nonvascularized) triggered potent indirect alloresponses. Altogether, these results suggest that primary vascularization of allografts is associated with a lack of indirect T cell alloreactivity. Finally, we show that long-term survival of vascularized skin allografts induced by anti-CD40L Abs was associated with a combined lack of indirect alloresponse and a shift of the direct alloresponse toward a type 2 cytokine (IL-4, IL-10)-secretion pattern but no activation/expansion of Foxp3(+) regulatory T cells. Therefore, primary vascularization of allografts governs their immunogenicity and tolerogenicity.

Novel effect of adenosine 5'-monophosphate on ameliorating hypertension and the metabolism of lipids and glucose in stroke-prone spontaneously hypertensive rats.

The aim of the study was to investigate the effects of adenosine 5'-monophosphate (AMP) in stroke-prone spontaneously hypertensive rats (SHRSP). Male rats (10 weeks old) were divided into three groups: a control group fed an AIN-93 M diet and two others fed supplemental AMP (17.5 and 87.5 mg/kg diet) for 3 weeks. AMP effectively improved hypertension, plasma triglyceride, and HDL-cholesterol, glucose, kidney function parameters, hepatic lipid, enhances plasma nitric oxide, and plasma adiponectin accompanied by the up-regulation of mRNA expression levels of the hepatic adiponectin receptor 2. Single and chronic oral administration of AMP affected the hepatic mRNA expression levels of genes involved in ß-oxidation, fatty acid synthesis, and AMP-activated protein kinase. Furthermore, a single oral dose of AMP (40 mg/kg body weight) improved hypertension and hyperglycemia in SHRSP. In conclusion, AMP displays a novel effect in ameliorating metabolic-related diseases in SHRSP and could be beneficial as a functional food.

Transplantation tolerance to a single noninherited MHC class I maternal alloantigen studied in a TCR-transgenic mouse model.

The mechanisms underlying tolerance to noninherited maternal Ags (NIMA) are not fully understood. In this study, we designed a double-transgenic model in which all the offspring's CD8(+) T cells corresponded to a single clone recognizing the K(b) MHC class I protein. In contrast, the mother and the father of the offspring differed by the expression of a single Ag, K(b), that served as NIMA. We investigated the influence of NIMA exposure on the offspring thymic T cell selection during ontogeny and on its peripheral T cell response during adulthood. We observed that anti-K(b) thymocytes were exposed to NIMA and became activated during fetal life but were not deleted. Strikingly, adult mice exposed to NIMA accepted permanently K(b+) heart allografts despite the presence of normal levels of anti-K(b) TCR transgenic T cells. Transplant tolerance was associated with a lack of a proinflammatory alloreactive T cell response and an activation/expansion of T cells producing IL-4 and IL-10. In addition, we observed that tolerance to NIMA K(b) was abrogated via depletion of CD4(+) but not CD8(+) T cells and could be transferred to naive nonexposed mice via adoptive transfer of CD4(+)CD25(high) T cell expressing Foxp3 isolated from NIMA mice.

Direct biliopancreatoduodenal invasion by hepatocellular carcinoma: report of the first resected case and review of the literature.

A rare resected case of hepatocellular carcinoma (HCC) invading the duodenum, the common bile duct (CBD), the gallbladder, and the pancreas is described. A 63-year-old man presented with a painful upper abdominal mass. Radiologic findings showed a 25-cm liver tumor arising from segment IV with an invasive extension to the hepatoduodenal ligament and pancreatoduodenal region, with a single intrahepatic metastasis. The patient successfully underwent a left hepatectomy in conjunction with a pylorus-preserving pancreatoduodenectomy (PD). As an unusual procedure, liver parenchymal transection was followed by PD to explore tumor resectability, because the overhanging liver mass precluded full hepatoduodenal ligament dissection. He was discharged without surgical complications, being free from antianalgesics, which had been used preoperatively. The main tumor was histologically diagnosed to be a poorly differentiated HCC with sarcomatous change invading the duodenum, the CBD, the gallbladder, and the pancreas. Unfortunately, aggressive hepatic and nodal recurrence, which was resistant to salvage chemotherapy, caused the patient's death at 8 months postoperatively. This is the first documented case of HCC with biliopancreatoduodenal invasion resected by hepatopancreatoduodenectomy. Literature review suggests a significant role of resection in selected patients with HCC with contiguous gastrointestinal tract invasion, particularly when the HCCs are naive without any forms of previous treatment. However, further surgical and nonsurgical experience is necessary to determine the oncological validity of aggressive surgery for HCC invading the biliopancreatoduodenal region.

Intracellular MHC class II controls regulatory tolerance to allogeneic transplants.

MHC class II (MHCII) genes have been implicated in the regulation of T lymphocyte responses. However, the mechanism of MHCII-driven regulation remains unknown. Matching for MHCII between donors and recipients of allografts favors regulatory T cell tolerance to transplants and provides a unique opportunity to study this regulation. In this study, we investigated MHCII regulation using transfer of donor MHCII genes in recipients of cardiac allografts. Transfer of MHCII IA(b) genes in the bone marrow of CBA mice (H-2(k)) prior to the grafting of IA(b+) fully allogeneic C57BL/6 (B6, H-2(b)) heart transplants resulted in donor-specific tolerance associated with long-term survival of B6, but not third-party, allografts without sustained immunosuppression. Strikingly, the majority of accepted heart transplants (>170 d) were devoid of allograft vasculopathy. Further studies indicated that intracellular IA(b) initiated the tolerogenic process, which was mediated by regulatory T cells (Tregs) that polarized antigraft responses to Th2 cytokine producers. This mechanism seems to be unique to MHCII genes, because previous MHC class I gene-based therapies failed to produce Tregs. These results demonstrate the key role of MHCII in the induction of Tregs. They also underscore a potential mechanism of specific inactivation of T cells in this model; when activated by IA(b+) grafts, IA(b)-specific Tregs repress the entire alloresponse to C57BL/6 transplants (including MHC I and minor Ags), thus mediating T cell tolerance.

Chemotherapy-induced sclerosing cholangitis as a rare indication for resection: report of a case.

Bile duct stricture due to chemotherapy-induced sclerosing cholangitis (CISC) is a potentially fatal complication of hepatic arterial infusion chemotherapy (HAIC). It is managed primarily with medical treatment and biliary stenting. We report a rare case of a CISC-related biliary stricture requiring resection. The patient had been receiving adjuvant HAIC for 11 months after a curative liver resection for hepatocellular carcinoma, when clinically overt cholangitis developed. Radiologic and biopsy findings suggested a CISC-related biliary stricture limited to the common hepatic duct. We discontinued HAIC and started corticosteroid treatment, which finally became ineffective. Endoscopic biliary stenting was impossible because of her severe biliary sclerosis, necessitating resection of the stricture, which was confirmed histologically to be secondary sclerosing cholangitis. The patient has shown no signs of recurrent cholangitis for 12 postoperative months since her operation. Thus, resection could be a treatment option for a CISC-related biliary stricture in selected patients.

Effects of immunosuppressants on induction of regulatory cells after intratracheal delivery of alloantigen.

BACKGROUND: We previously reported that intratracheal delivery (ITD) of alloantigen generated regulatory cells in mice. Here, we examined the effect of various doses of conventional immunosuppressants (FK506, cyclosporine A, azathioprine, mycophenolate mofetil, and rapamycin) on inducing regulatory cells in our model. METHODS: CBA mice (primary recipients) were given C57BL/6 splenocytes by ITD and either no additional treatment or various doses of an immunosuppressant. Seven days later, splenocytes from these mice were adoptively transferred into naive secondary CBA recipients that underwent C57BL/6 cardiac grafting the same day. RESULTS: Adoptive transfer from primary recipients given ITD of splenocytes alone induced prolonged allograft survival in secondary recipients (median survival time [MST], 50 days), suggesting that regulatory cells were generated. When ITD of alloantigen was combined with daily administration of 0.1 mg/kg FK506 or 0.2 mg/kg rapamycin, graft survival was similarly prolonged (MST 55 and 50 days, respectively). When combined with 20 or 40 mg/kg MMF or 0.4 mg/kg rapamycin, the majority of recipients demonstrated indefinite survival (MST, >100 days in all groups). When ITD of alloantigen was combined with 0.3, 0.5, or 1.0 mg/kg FK506; 5, 10, or 25 mg/kg cyclosporine A; or 1.0 or 2.0 mg/kg azathioprine, allografts were rejected acutely (MST 7-13 days). CONCLUSION: Generation of regulatory cells by ITD of alloantigen was facilitated by mycophenolate mofetil and high doses of rapamycin but abrogated by cyclosporine A, azathioprine, and high doses of FK506. Low doses of rapamycin and of FK506 did not interfere with generation of regulatory cells.

Graft-versus-host-reactive donor CD4 cells can induce T cell-mediated rejection of the donor marrow in mixed allogeneic chimeras prepared with nonmyeloablative conditioning.

Murine mixed hematopoietic chimerism can be achieved following nonmyeloablative conditioning with cyclophosphamide, T cell-depleting monoclonal antibodies, and thymic irradiation. Donor lymphocyte infusions (DLIs) 35 days after bone marrow transplantation (BMT) convert mixed to full donor chimerism and mediate graft-versus-lymphoma effects without graft-versus-host disease. We evaluated the role of T-cell subsets in DLIs in converting mixed to full donor chimerism in a fully major histocompatibility complex-mismatched strain combination. Whereas DLIs administered on day 35 converted 100% of mixed chimeras to full donor chimerism, conversion was less frequent when either CD4 or CD8 cells were depleted, indicating that both subsets contribute to the conversion. Surprisingly, administration of CD8-depleted DLIs led to complete loss of donor chimerism in a high proportion (54%) of recipients compared with CD4-plus CD8-depleted DLIs (15%) or CD4-depleted DLIs (0%) (P <.05). DLIs administered at early time points after BMT (eg, day 21) also precipitated rejection of donor marrow by recipient alphabeta T cells, in association with donor CD4 cell expansion and high production of interleukin 2 (IL-2), IL-4, and interferon-gamma. Thus, DLIs can paradoxically induce marrow rejection by residual host alphabeta T cells. These results have implications for the timing of and use of subset depletion of DLIs in recipients of nonmyeloablative transplants.

Induction of operational tolerance and generation of regulatory cells after intratracheal delivery of alloantigen combined with nondepleting anti-CD4 monoclonal antibody.

BACKGROUND: We previously showed that intratracheal delivery of alloantigen induced prolonged survival of fully allogeneic cardiac grafts in mice. Here, this treatment protocol was combined with nondepleting anti-CD4 monoclonal antibody (mAb) to induce operational tolerance. METHODS: CBA (H-2k) mice were pretreated with intratracheal delivery of whole splenocytes from C57BL/10 (H-2b) mice or a 15-mer Kb peptide, with or without intraperitoneal administration of nondepleting anti-CD4 mAb (YTS177). Seven days later, C57BL/10 hearts were transplanted into the pretreated CBA mice. In addition, some naive CBA mice underwent adoptive transfer of splenocytes from pretreated CBA mice and transplantation of a C57BL/10 heart on the same day. RESULTS: Untreated CBA mice rejected C57BL/10 cardiac grafts acutely (median survival time, 12 days). Mice given intratracheal delivery of whole splenocytes or Kb peptide demonstrated prolonged graft survival (median survival time, 84 and 76 days, respectively). Concurrent administration of YTS177 and intratracheal delivery of splenocytes or Kb peptide resulted in indefinite graft survival. Mice with long-surviving C57BL/10 cardiac grafts showed acceptance of skin grafts from C57BL/10 mice but not BALB/c mice, demonstrating that operational tolerance had been induced. Adoptive transfer of splenocytes from mice pretreated with intratracheal delivery of splenocytes or Kb peptide plus YTS177 induced indefinite survival of cardiac grafts in secondary recipients, indicating that regulatory cells had been generated. CONCLUSION: In a murine model, intratracheal delivery of donor splenocytes or Kb peptide combined with YTS177 induced operational tolerance and generated regulatory cells.

CD27/CD70, CD134/CD134 ligand, and CD30/CD153 pathways are independently essential for generation of regulatory cells after intratracheal delivery of alloantigen.

BACKGROUND: We investigated whether blockade of tumor necrosis factor receptor-ligand pathways could generate regulatory cells induced by intratracheal delivery of alloantigen. METHODS: CBA (H-2k) mice were pretreated with intratracheal delivery of splenocytes (1x10(7)) from C57BL/10 (H-2b) mice and intraperitoneal administration of monoclonal antibody (mAb) specific for CD70, CD134 ligand (CD134L), CD153, or CD137L. Seven days later, C57BL/10 hearts were transplanted into pretreated CBA mice. Some naive CBA mice underwent adoptive transfer of splenocytes (5x10(7)) from pretreated CBA mice and transplantation of a C57BL/10 heart on the same day. RESULTS: Untreated CBA mice rejected C57BL/10 cardiac grafts acutely (median survival time [MST] 12 days). Pretreatment with intratracheal delivery of C57BL/10 donor splenocytes prolonged graft survival significantly (MST 84 days). Mice given intratracheal delivery of alloantigen plus anti-CD70, anti-CD134L, or anti-CD153 mAb, but not those given intratracheal delivery of alloantigen plus anti-CD137L mAb, rejected their graft acutely (MST 16, 14, 10, and 65 days, respectively). Adoptive transfer of splenocytes from mice pretreated with intratracheal delivery of alloantigen plus anti-CD70, CD134L, or CD153 mAb did not prolong survival of C57BL/10 cardiac grafts in naive secondary CBA recipients (MST 14, 11, and 11 days, respectively), whereas adoptive transfer of splenocytes from mice given intratracheal delivery of alloantigen plus anti-CD137L mAb did (MST 75 days). CONCLUSION: The CD27/CD70, CD134/CD134L, and CD30/CD153 pathways are independently required for generation of regulatory cells in our model.

Role of the CTLA4 pathway in hyporesponsiveness induced by intratracheal delivery of alloantigen.

BACKGROUND: The authors previously reported that intratracheal delivery (ITD) of donor alloantigen induced donor-specific hyporesponsiveness to C57BL/10 cardiac allografts in CBA recipients and that blockade of the B7 pathways abrogated that hyporesponsiveness. In this study, the authors used a CD28-deficient model to evaluate which signal, either through CD28 or cytotoxic T-lymphocyte-associated antigen (CTLA4), is involved in the induction of hyporesponsiveness. METHODS: Seven days before transplantation of hearts from C3H/HeJ (H2k) mice into C57BL/6 (H2b) or CD28-deficient (C57BL/6 background) mice, the transplant recipients were given ITD of donor splenocytes (1 x 10(7)), alone or in combination with human CTLA4-immunoglobulin (Ig) (200 microg). RESULTS: ITD of C3H splenocytes induced donor-specific hyporesponsiveness to C3H cardiac grafts in C57BL/6 recipients (graft median survival time [MST], 40 days). Administration of CTLA4-Ig concurrently with ITD abrogated the prolonged allograft survival (MST, 12 days). Interestingly, ITD of C3H splenocytes induced prolonged survival of C3H allografts in CD28-deficient recipients (MST, 55 days). Furthermore, administration of CTLA4-Ig combined with ITD of C3H splenocytes abrogated the prolonged survival of C3H allografts in CD28-deficient recipients (MST, 7 days), whereas recipients given isotype-control antibody in combination with ITD of splenocytes had prolonged survival of C3H allografts (MST, 58 days). CONCLUSIONS: Taken together, the authors' findings indicate that a signal through CTLA4, rather than through CD28, plays an important role in the induction of hyporesponsiveness by ITD of alloantigen in this model.

Induction of indefinite survival of fully allogeneic cardiac grafts and generation of regulatory cells by intratracheal delivery of alloantigens under blockade of the CD40 pathway.

BACKGROUND: The authors previously showed that intratracheal delivery (ITD) of donor splenocytes induced prolonged survival of fully allogeneic cardiac grafts in mice. In this study, this treatment protocol was combined with blockade of the CD40 pathway in an attempt to induce operational tolerance. METHODS: CBA mice were given donor splenocytes (1x107) or Kb peptide (100 microg) by ITD with or without antibody specific for mouse CD40 ligand (MR1, 200 microg) 7 days before transplantation of a C57BL/10 heart. Also, splenocyte (5 x 107) from primary recipient CBA mice given ITD of donor splenocytes or Kb peptide plus MR1 were adoptively transferred into naive CBA secondary recipients 7 days after the pretreatment and C57BL/10 hearts were transplanted into those recipients the same day. RESULTS: ITD of donor splenocytes and Kb peptide induced prolonged survival of cardiac grafts (median survival time [MST], 74 and 56 days, respectively), whereas naive control mice and mice pretreated with syngeneic splenocytes had acute graft rejection (MST in both groups, 7 days). When MR1 was included, all grafts survived indefinitely (>200 days), but mice pretreated with MR1 alone had graft rejection (MST, 54 days). Mice bearing cardiac grafts had acceptance of skin grafts from C57BL/10 but not BALB/c mice, demonstrating that operational tolerance was induced. Secondary recipients given adoptive transfer of splenocytes from primary recipients of the combined treatment had acceptance of C57BL/10 grafts, suggesting that regulatory cells were generated within 7 days of pretreatment. CONCLUSIONS: ITD of donor splenocytes or Kb peptide under blockade of the CD40 pathway induced operational tolerance and generated regulatory cells.

High dose of antithrombin III induces indefinite survival of fully allogeneic cardiac grafts and generates regulatory cells.

BACKGROUND: The authors investigated whether antithrombin III (AT-III) could induce unresponsiveness to alloantigens. METHODS: CBA mice were given intravenous injection of 50 or 500 U/kg AT-III or control plasma the same day as transplantation of a heart from a C57BL/6 mouse. An adoptive transfer study and mixed leukocyte culture analysis were also performed. RESULTS: Naive CBA mice rejected C57BL/6 cardiac grafts acutely (median survival time [MST], 9 days). The 50-U/kg dose of AT-III induced a moderate increase in graft survival (MST, 25 days), whereas control mice rejected their graft acutely (MST, 7 days). With the 500-U/kg dose of AT-III, all grafts survived indefinitely (>100 days) and regulatory cells were generated. In vitro, AT-III suppressed proliferation of mixed leukocyte responses and generation of interleukin-2. CONCLUSION: AT-III can be not only an antithrombotic agent but also a strong immunomodulating agent when used at high dose.

Intratracheal delivery of a single major histocompatibility complex class I peptide induced prolonged survival of fully allogeneic cardiac grafts and generated regulatory cells.

We have previously reported that intratracheal delivery of donor splenocytes in mice induces hyporesponsiveness to fully allogeneic cardiac grafts and generates regulatory cells. Here, we examined whether an allopeptide would produce the same results. A 15-mer (54-68) peptide corresponding to a hypervariable region of the K(b) molecule was given intratracheally or intravenously to CBA (H2(k)) mice 7 days before transplantation of a C57BL/10 (H2(b)) or BALB/c (H2(d)) heart and was also used in adoptive transfer experiments. Cardiac grafts in recipients given K(b) peptide intratracheally experienced a median survival time (MST) of 56 days, whereas those in recipients given the peptide intravenously were rejected acutely (MST=7.5 days). Adoptive transfer of splenocytes from mice pretreated intratracheally with K(b) peptide to naïve secondary recipients prolonged survival of cardiac grafts (MST = 35 days). Intratracheal delivery of a single major histocompatibility complex class I peptide induced hyporesponsiveness to allogeneic cardiac grafts and generated regulatory cells.

B7/CTLA4 pathway is essential for generating regulatory cells after intratracheal delivery of alloantigen in mice.

BACKGROUND: The mechanism of hyporesponsiveness induced by intratracheal (IT) delivery of alloantigen was examined and its effect on cardiac graft survival was assessed in studies in mice. METHODS: In CBA (H2 ) mice, donor splenocytes were given by IT delivery 7 days before transplantation of a C57BL/10 (H2 ) heart. To determine whether regulatory cells were involved in hyporesponsiveness, splenocytes from mice given IT delivery of alloantigen and antibodies for B7-1, B7-2, or CTLA4 were adoptively transferred to naïve secondary recipients 7 days after delivery; those recipients underwent heart transplantation the same day. Effects on cell proliferation and cytokine production of splenocytes from mice given IT delivery of alloantigen were examined in mixed leukocyte cultures (MLC). RESULTS: Cardiac graft survival was significantly prolonged in mice given IT delivery of alloantigen (median survival time [MST], 81 days); those given syngeneic splenocytes rejected grafts acutely (MST, 7 days; P<0.05). Adoptive transfer of splenocytes also significantly prolonged survival of cardiac grafts in secondary recipients (MST, 62 days). When B7-1, B7-2, or CTLA4 antibody was combined with IT delivery of alloantigen in the first recipient, all grafts were rejected within 14 days in second recipients after adoptive transfer. In mixed leukocyte cultures, splenocytes from these mice did not respond to alloantigen and production of interleukin-4 and interleukin-10 was increased. CONCLUSIONS: Donor splenocytes delivered IT induced hyporesponsiveness and regulatory cells in our animal model, and such induction was dependent on B7-1, B7-2, and CTLA4 signals.