Initial burst of viremia related to CD8 effector memory T cells after living donor liver transplantation in hepatitis C virus-infected recipients.

The post-transplant immune responses, viremia, and allograft histology after living donor liver transplantation were studied in 39 hepatitis C virus (HCV)-infected recipients. The recipients were classified into the following groups according to a hierarchical clustering of their preoperative CD8CD45 T-cell isoforms: group I, naive-dominant; group II, effector memory-dominant; and group III, effector-dominant. Plasma HCV-RNA rapidly increased and then peaked as an initial burst around postoperative day (POD) 25 in group I, on POD 40 in group II, and on POD 55 in group III. The initial burst of viremia was suppressed by the high expression of CD8+CD28-CD27- subsets. The progression of fibrosis > or =F2 was significantly more frequent for those patients with the highest viremia levels. Moreover, the initial T-cell immune response became less important throughout time, and new immune responses emerged after 2 months that modified the host-virus interaction. It is suggested that the interferon (IFN)-alpha/ribavirin therapy starting 2 months may be an effective option and now is undertaken.

Impact of preexisting circulating effector T cells on the outcome of ABO-incompatible adult LDLT.

Our aim was to clarify the significance of phenotype of circulating CD8 T(+) cells on the outcome of ABO-incompatible (ABO-I) living donor liver transplantation (LDLT). Twenty-six recipients undergoing ABO-I LDLT and 92 undergoing ABO-compatible (ABO-C) LDLT were classified into three groups according to preoperative proportion of CD8 T(+) cells: naive-dominant (group I), effector memory-dominant (group II), and effector-dominant (group III) recipients. The clinical courses were analyzed. The results showed that in ABO-C groups I and II and in ABO-I group I, effector cells remained above the pretransplant levels after tacrolimus administration. However, in ABO-C group III and ABO-I groups II and III, effector cells were down-regulated for a prolonged period, along with markedly decreased perforin expression and frequent life-threatening complications. ABO-I group II and group III recipients had higher infection rates. It was concluded that recipients with preexisting high effector CD8 T(+) cells are unfavorable candidates for ABO-I LDLT.

Serial assessment of immune status by circulating CD8 effector T cell frequencies for posttransplant infectious complications.

To clarify the role of CD8+ effector T cells for infectious complications, 92 recipients were classified according to the hierarchical clustering of preoperative CD8+CD45 isoforms: Group I was naive, Group II was effector memory, and Group III was effector (E) T cell-dominant. The posttransplant infection rates progressively increased from 29% in Group I to 64.3% in Group III recipients. The posttransplant immune status was compared with the pretransplant status, based on the measure (% difference) and its graphical form (scatter plot). In Groups I and II, both approaches showed a strong upward deviation from pretransplant status upon posttransplant infection, indicating an enhanced clearance of pathogens. In Group III, in contrast, both approaches showed a clear downward deviation from preoperative status, indicating deficient cytotoxicity. The % E difference and scatter plot can be used as a useful indicator of a posttransplant infectious complication.

Current status of liver transplantation across ABO blood-type barrier.

Outcomes of ABO-blood type incompatible liver transplantation have recently improved owing to various treatments. The typical clinical manifestations of antibody mediated rejection (AMR) are hepatic necrosis and intrahepatic biliary complication (IHBC). The prognosis of AMR is poor. AMR is the result of circulatory disturbance which is caused by injury to the endothelium due to an antibody-antigen-complement reaction. Diffuse C4d staining in the portal capillaries and periportal areas in severe AMR. Since natural antibodies against A/B carbohydrate determinants are likely to develop as a result of exposure to environmental bacteria that express similar determinants, the B-1 lineage has been speculated to be the major population of B-cell types responding to A/B determinants. Calcineurin inhibitors block B-1 cell differentiation. Rituximab can be used to deplete both cells that are producing IgM antibodies and those that have already differentiated into B-1 cells. Mycophenolate mofetil is required to inhibit the production of IgG subclass of antibodies. The outcome is now similar to that of blood-type-matched transplantation. However, there are still issues to be solved in order to further improve the outcome via a decrease of infection.

Initial steroid bolus injection promotes vigorous CD8+ alloreactive responses toward early graft acceptance immediately after liver transplantation in humans.

We have found that steroid bolus withdrawal prior to graft reperfusion increased the incidence of acute cellular rejection (ACR). This study aims to clarify how initial steroid bolus (ISB) injection at reperfusion influences the kinetics of CD8(+) alloreactive immune responses immediately after living donor liver transplantation (LDLT). A total of 49 hepatitis C virus (HCV)-infected recipients were classified into 3 groups according to hierarchical clustering by preoperative CD8(+)CD45 isoforms. The naive T cell proportion was considerably higher in Group I than in Groups II and III, whereas Group II recipients had the highest effector memory (EM) T cells and Group III the highest effector T cells. The frequency of ACR was significantly higher in recipients without ISB than in those with ISB. In particular, the ACR rates were the highest in Group II without ISB. Following ISB, the proportion of effector T cells was promptly upregulated within 6 hours after graft reperfusion, simultaneously with the upregulation of CD27(-)CD28(-) subsets, interferon-gamma (IFN-gamma), tumor necrosis factor-alpha and perforin expression, which significantly correlated with increasing interleukin (IL)-12 receptor beta 1 cells. These were then downregulated to below preoperative levels by tacrolimus (Tac) administered at 24 hours. These changes did not occur in the absence of ISB. In Group II without ISB, the downregulation of IL-12Rbeta1(+) cells was the greatest, consistent with the highest rates of ACR and mortality (60%). In conclusion, ISB must be done in place, especially in Group II with preexisting high EM T cells, to enable the development of early allograft acceptance.

B-cell surface marker analysis for improvement of rituximab prophylaxis in ABO-incompatible adult living donor liver transplantation.

Although the effectiveness of rituximab has been reported in ABO blood group (ABO)-incompatible (ABO-I) organ transplantation, the protocol is not yet established. We studied the impact of the timing of rituximab prophylaxis and the humoral immune response of patients undergoing ABO-I living donor liver transplantation (LDLT), focusing on clinicopathological findings and the B-cell subset. From July 2003 to December 2005, 30 adult patients were treated with hepatic artery infusion (HAI) protocol without splenectomy for ABO-I LDLT. A total of 17 patients were treated only with HAI (no prophylaxis), and the other 13 were treated with rituximab prophylaxis at various times prior to transplantation. For B-cell study of the spleen, another 4 patients undergoing ABO-I LDLT both with HAI after prophylaxis and eventual splenectomy, and 3 patients with ABO-compatible LDLT with splenectomy were enrolled. The mortality of the 30 patients with HAI, without splenectomy, and with/without rituximab prophylaxis was 33% and the main cause of death was sepsis. Peripheral blood B cells were completely depleted, anti-donor blood-type antibody titer was lower, and clinical and pathological antibody-mediated rejection was not observed in patients with prophylaxis earlier than 7 days before transplantation (early prophylaxis). Early rituximab prophylaxis significantly depleted B cells and memory B cells in the spleen but not in lymph nodes. On the other hand, B cells and memory B cells increased and memory B cells became dominant during antibody-mediated rejection. In conclusion, early prophylaxis with rituximab depletes B cells, including memory B cells, in the spleen and is associated with a trend toward lower humoral rejection rates and lower peak immunoglobulin (Ig)G titers in ABO-I LDLT patients.

Cytotoxic T-cell-mediated defense against infections in human liver transplant recipients.

Previous studies have shown that postoperative infection is highest in transplant recipients with preexisting high levels of cytotoxic T lymphocytes (CTLs). To study this phenomenon, 106 adult liver transplant recipients were divided into 3 groups, based on hierarchical clustering of the CD3(+)CD8(+)CD45 isoform fractions prior to living donor liver transplantation (LDLT). Group I had the highest naive T-cell levels (subset CD45RO(-)CCR7(+)), Group II had the highest effector/memory (EM) T-cell levels (subset CD45RO(+)CCR7(-)), and Group III had the highest effector T-cell levels (subset CD45RO(-)CCR7(-)). In Group I, CTLs upregulated in response to invading pathogens much earlier and more rapidly than the other groups; this response was associated with CD4(+) T-cell help, downregulation of CD27(+)CD28(+) subsets, and upregulation of interferon-gamma and perforin expression. In contrast, in Groups II and III, CTLs upregulated slowly following persistent viral infection and did not respond efficiently to acute infection. In addition, Group II's cytolytic responses were due mainly to upregulation of the CD8(+) EM T-cell fraction, whereas Group III's cytolytic responses were attributable to upregulation of effector T cells. The prevalence of EM or effector T cells was dependent on differentiation of the CD8(+) phenotype before LDLT. In conclusion, in most infected transplant recipients who died, generation of CD8(+) CTLs had been suppressed without associated CD4(+) T-cell help.

[A randomized controlled study comparing uracil-tegafur (UFT)+tamoxifen (UFT+TAM therapy) with cyclophosphamide+adriamycin+5-fluorouracil (CAF therapy) for women with stage I , II, or IIIa breast cancer with four or more involved nodes in the adjuvant setting].

We performed a controlled study to compare the response to cyclophosphamide (CPA), adriamycin (ADM), and fluorouracil (5-FU) (CAF therapy) with that to uracil-tegafur (UFT) plus tamoxifen (TAM) (UFT+TAM therapy), when given as postoperative adjuvant therapy to women with breast cancer. The patients were registered from September 1991 through February 1995 at 51 institutions in the Kinki district of Japan. All patients had stage I, II, or IIIa breast cancer with four or more lymph-node metastases and underwent mastectomy. CAF therapy and UFT+TAM therapy were started within 4 weeks after surgery. CAF therapy consisted of CPA (100 mg/day) on days 1 to 14, followed by 2 weeks of rest, plus ADM (20 mg/m(2)/day) on days 1 and 8 and 5-FU (300 mg/m(2)/day) on days 1 and 8. A total of 6 courses were delivered. UFT+TAM therapy consisted of 3 years of UFT (400 mg/day) plus TAM (20 mg/day), given daily. CAF therapy and UFT+TAM therapy were each assigned to 82 patients. The 5-year survival rate was significantly higher in the UFT+TAM group (82.1%) than in the CAF group (66.2%; p=0.04, logrank test). The 5-year relapse-free survival rate was higher in the UFT+TAM group (61.8%) than in the CAF group (46.3%; p=0.07, logrank test). As for adverse events, the rates of leukopenia, anorexia, nausea and vomiting, general malaise, and hair loss were lower in the UFT+TAM group than in the CAF group. These results suggest that long-term treatment with UFT+TAM may be a useful alternative adjuvant therapy for the management of breast cancer, especially in elderly patients.

Classification of human liver transplant recipients by their preoperative CD8+ T cell subpopulation and its relation to outcome.

The primed status of T cells is markedly different among liver transplant recipients, due to a lifetime of antigen exposure and reduced thymopoiesis by aging, and diseases. This study aims to characterize the preoperative immunological status of CD8+ T cell subpopulations and relate it to the outcome for liver transplant recipients. We classified 112 liver transplant recipients into 5 groups, based on hierarchical clustering of the CD8+CD45 isoform proportion of T cells. In Groups I and II (pediatric), the naive T cell proportion was more than 50%. In adult recipients, Group III was characterized by a naive T cell proportion of 50%, Group IV had the greatest effector/memory T cells (EM), and Group V had the greatest proportion of effector T cells. In Groups IV and V, the effector T cell proportion was considerably higher, and was accompanied by marked downregulation of the CD27+CD28+ subsets and upregulation of interferon gamma (IFN)-gamma, tumor necrosis factor-alpha, and perforin expression. Group V recipients tended to be complicated postoperatively, with a significantly reduced survival rate (1 yr, 66.8%) and markedly reduced Eastern Cooperative Oncology Group performance status.

Role of bcl-2 mRNA in homeostatic proliferation in circulating T-cells in human liver transplant patients after T-cell depletion.

BACKGROUND: Prolonged T-cell depletion after liver transplantation leads to life-threatening infections. Members of the anti-apoptotic Bcl-2 gene family can maintain T-cell viability. T-cell numbers and their Bcl-2 expression following living donor liver transplantation (LDLT) were analyzed in 108 surviving and 13 deceased recipients. MATERIALS AND METHODS: Bcl-2 mRNA levels and phenotypic changes of T-cells were examined by quantitative PCR and by measuring expression of CD45RO and CCR7. RESULTS: Based on the restoration of peripheral T-cell numbers, the 108 surviving recipients were classified into three groups. All recipients showed T-cell depletion, down to approximately 30% of pretransplant levels within 3 h of graft reperfusion. In Group I, the T-cell numbers were rapidly restored to pretransplant levels, within 5 days, with a rapid decrease in Bcl-2 mRNA levels immediately after LDLT. In Group II, the T-cell numbers were restored to normal levels by 19 days, with down-regulation of Bcl-2 mRNA. In Group III, the T-cell numbers were maintained at low levels for much longer, with high levels of Bcl-2 mRNA. In all three groups of recipients, there was statistically significant (r = -0.78) inverse correlation between T-cell numbers and Bcl-2 mRNA. CONCLUSIONS: For successful transplantation, homeostatic restoration of T-cells must occur as soon as possible. Evaluation of peripheral T-cell numbers and of Bcl-2 expression may have therapeutic potential in identifying those transplant patients who face increased risk of infection.

Quantitative PCR to evaluate small amounts of BCL2 mRNA in human peripheral T cells: implication of equimolar target and competitor end products.

BACKGROUND: It is difficult to reliably and reproducibly quantitate small amounts of mRNA by conventional PCR. The method we describe here enabled us to more accurately quantitate a small amount of BCL2 mRNA in human peripheral T cells. METHODS: The sample was amplified in four tubes containing three-fold serial dilutions of competitor so that when the PCR products in the four tubes were totaled, the number of target and competitor components were approximately equal. An unknown concentration of target molecules should be assessed only within a narrow range, requiring that several reactions be performed for each sample. RESULTS AND CONCLUSION: With this method, conventional PCR machines can be used to perform quantitative PCR on very small amounts of BCL2, and would be especially useful for samples that contain substance(s) that affect PCR amplification efficiency.