Alterations in T lymphocyte subpopulations associated with renal allograft rejection

The peripheral blood OKT3 (total T), OKT4 (T helper/inducer), and OKT8 (T suppressor/cytotoxic) cells were determined by flow cytometry on twenty consecutive recipients of HLA-nonidentical cadaveric renal allografts. The absolute number of cells in all three populations decreased significantly posttransplantation, but no differences were found between patients experiencing rejection and those in quiescence. An OKT4/OKT8 ratio of greater than or equal to 1.7, either pretransplant or posttransplant, uniformly identified patients who subsequently experienced rejection. However, an OKT4/OKT8 ratio of less than 1.7 did not identify patients with a low risk of rejection. Pretransplant splenectomy was performed in 6 of 7 patients who rejected despite a low ratio. Serial monitoring of the OKT4/OKT8 ratio posttransplantation determined that an increase in the ratio of greater than or equal to 0.5 was a sensitive (81%) and specific (91%) indicator of a rejection episode. Graft survival was improved in patients with a high posttransplant OKT4/OKT8 ratio. These results indicate that the balance of helper and suppressor cell function may be of critical importance to the fate of an allograft, and that the alterations in this balance can be used to assist in the clinical management of allograft recipients.     

Superior 3-year kidney graft function in patients with impaired pretransplant Th2 responses

Abstract In a prospective study of 80 patients, we investigated the association of kidney graft rejection with pretransplant CD4 helpersuppressor function, B cell responses, and in vitro cytokine secretion. A pokeweed mitogen-driven allogeneic coculture system was used to assess CD4 helper/suppressor function and T cell-dependent B cell responses. SAC I was used for T celland monocyte-independent stimulation of B cell cultures. B cell differentiation was assessed in a reverse hemolytic plaque assay. ELISAs were used to determine in vitro cytokine secretion. None of the 12 patients with pretransplant CD4 helper defects (<10% helper activity) had acute rejection episodes in contrast to 32 of 68 (47%) patients with normal pretransplant CD4 helper function ( P = 0.001). Patients with pretransplant CD4 helper defects exhibited better 3-year graft function than patients without CD4 helper defects (serum creatinine of functioning grafts: 1.2 ± 0.1 mg/dl compared to 1.7 ± 0.1 mg/dl, P < 0.05). Low pretransplant IL-10 responses (<100 pg/ml; 14/80 patients) were significantly associated with a low incidence of acute rejection episodes ( P < 0.01) and good 3- year graft function ( P < 0.05). These data show that impaired pretransplant Th2 responses-CD4 help and IL-10 responses-predict a low risk of kidney graft rejection and good 3-year graft function, whereas Th1 (IL-2, IFN-γ) and B cell/monocyte responses are not of predictive value.     

Requirements for the induction of allospecific CD8+ suppressor T cells in the rat primary mixed lymphocyte response. CD4+, CD45R+ T cells, or supernatant factor

We examined the requirements for the induction of the MLR-generated allospecific CD8+ suppressor T cells in the rat. Depleting the responder population of CD4+ T cells before initiating the primary MLR abrogates the generation of day-5 CD8+ T suppressor effectors. Readdition of at least 10% CD4+ T cells to the CD4+ depleted primary MLR reconstitutes suppressor cell generation. Using the anti-CD45R monoclonal antibody OX22, we also show that the T suppressor inducer cells are CD4+ CD45R+. Using a dual chamber Transwell culture system, which allows cells to be co-incubated without direct cell-to-cell contact, we show that a soluble factor/s, produced during the course of the primary MLR, is capable of inducing naive CD8+ T cells to become suppressor effectors but only when these CD8 T cells are in direct contact with allogeneic stimulators. Allospecificity is conferred by the stimulator cells and not by the suppressor-inducer factor. The supernatant of day-5 primary MLR is also capable of inducing antigen-specific suppressor effectors from naive CD8+ T cells, and also only in the presence of allogeneic stimulator cells. Recombinant human IL-2, in doses that are up to five times the amount present in the supernatant cultures, is unable to induce suppressor-effector cells from naive CD8+ T cells. We conclude that, to become allospecific suppressor effectors, naive CD8+ T cells require contact with allogeneic stimulator cells and either CD4+ CD45R+ suppressor inducer cells or suppressor inducer factor/s produced during the course of the primary MLR.     

Pretransplant Numbers of CD16+ Monocytes as a Novel Biomarker to Predict Acute Rejection After Kidney Transplantation: A Pilot Study

Acute rejection is one of the major immunological determinants of kidney graft function and survival. Early biomarkers to predict rejection are lacking. Emerging evidence reveals a crucial role for the monocyte/macrophage lineage cells in the pathogenesis of rejection. We hypothesized that higher pretransplant numbers of proinflammatory CD16+ monocytes can predict rejection. The study cohort consisted of 104 kidney transplant recipients (58 with no rejection and 46 with biopsy‐proven rejection) and 33 healthy persons. Posttransplant median follow‐up time was 14.7 mo (interquartile range 0.3–34 mo). Pretransplantation blood samples were analyzed by flow cytometry for monocyte immunophenotypes. Groups were compared by Cox regression models for the occurrence of acute rejection. We documented a significantly increased absolute number of pretransplant CD16+ monocytes in patients who developed biopsy‐proven rejection after transplantation compared with those with no rejection (hazard ratio [HR] 1.60, 95% CI 1.28–2.00, p < 0.001) and healthy persons (HR 1.47, 95% CI 1.18–1.82, p < 0.001). In parallel, significantly fewer absolute numbers of CD16? monocytes were observed at pretransplant time points in rejectors versus nonrejectors (HR 0.74, 95% CI 0.58–0.94, p < 0,014). A higher pretransplant number of CD16+ monocytes is significantly associated with a higher risk of acute rejection after kidney transplantation.     

Effect of hormone replacement therapy on CD4+ and CD8+ numbers, CD4+/CD8+ ratio, and immunoglobulin levels in hemodialysis patients

Uremia induces a suppression of the immune status. A large clinical literature suggests that estradiol (E2) plays a critical role in immune function. A large proportion of women hemodialysis patients faced early menopause and inadequate estrogen levels. The aim of the present study is to evaluate the effect of hormone replacement therapy on immune function in terms of CD4+ numbers (inducer/helper T cells), CD8+ numbers (cytotoxic/suppressor T cells), CD4+ / CD8+ ratio, and IgG, IgM, IgA levels in woman hemodialysis patients. In our study, 15 female hemodialysis patients (median age 32.6 range 24-45) were treated with triphasic estrogen/progesterone preparation (estradiol 2 mg for 10 days, and afterwards estradiol 2 mg+norethisterone 1 mg for another 10 days, and at the end estradiol 1 mg for 6 days) for 6 months. CD4+ numbers, CD8+ numbers, and IgG, IgA, and IgM levels were determined before and after HRT. The "paired-samples T" test was used for statistical analysis of pretreatment and posttreatment values. A significant increase was observed for CD4+ numbers (582 +/- 435 versus 637 +/- 445, p = 0.04) and CD4+/CD8+ ratio (1.4 +/- 0.16 to 2.4 +/- 0.3, p < 0.01) after hormone replacement therapy (HRT). Serum immunoglobulin levels were not changed significantly. In conclusion, in postmenopausal hemodialysis patients, HRT significantly increased CD4+ numbers and CD4+ / CD8+ ratio, but no effect was observed in IgM, IgG, and IgA levels. Long-term clinical effects of HRT on immune system should be investigated in dialysis patients with further studies.     

Cellular infiltrate in renal graft rejection: T lymphocyte subsets detected by monoclonal antibodies

We have examined the interstitial cellular infiltrate using monoclonal antibodies against T cells (Cris 1), helper/inducer T cells (OKT4) and suppressor/cytotoxic T cells (OKT8) by indirect immunofluorescence in renal biopsies taken from 14 transplanted patients during clinical episodes suggestive of acute (n = 9), chronic (n = 2) and no rejection (n = 3). Infiltrating T cells and T cell subsets were found to be significantly increased during all types of rejection (n = 11) as compared to no rejection (n = 3). Two types of biopsies could be distinguished according to the predominance of T cell subsets. In some biopsies (n = 6), OKT8+ cells were significantly more numerous that OKT4+ cells. In the remaining biopsies (n = 5), OKT4+ cells were more common that OKT8+ cells, the OKT4/OKT8 ratio being significantly higher. No association was observed between HLA mismatch and predominating T cell subset, neither for type nor outcome of graft rejection. Our results suggest that the OKT4+ cells may play a more important role than previously reported in renal graft rejection.     

Identification of the cellular subpopulations infiltrating rejecting cadaver renal allografts. Preponderance of the T4 subset of T cells

In the rejection response against renal allografts, the relative importance of helper/inducer T cells mediating a delayed-type hypersensitivity response and of T cells with direct cytotoxicity has not been defined. These subpopulations were identified with commercially available monoclonal antibodies and an indirect immunoperoxidase technique in 31 renal biopsies from patients undergoing acute rejection episodes and in 9 rejected nephrectomy specimens. T lymphocytes were the predominant cell population in all biopsies and in 8 of 9 nephrectomies. The T4 helper/inducer subset was equal to, or greater than, the T8 cytotoxic/suppressor subset in 28 of the 31 biopsies and in the 8 nephrectomy specimens that had histological evidence of cellular rejection. T4 lymphocytes were found predominantly in large areas of cellular infiltrate. T8 lymphocytes had a more diffuse interstitial distribution and were a minority of the cells in the large areas of cellular infiltration. These results show that helper/inducer T lymphocytes are often more frequent than cytotoxic/suppressor cells in acute renal allograft rejection in humans and they suggest that helper/inducer T cells may play an important role in the mediation of graft destruction.     

Impairment of helper T-cell function following severe head injury.

Major infections, such as sepsis and pneumonia, occur in 50-75% of patients following isolated severe head injury. Previous studies have demonstrated that this high incidence of infection following severe head injury may be related to a decrease in helper T-cell activation and function. The present study was designed to investigate the effect of severe head injury on specific subgroups of helper T cells known to enhance or suppress cellular immune function. Specifically, peripheral blood lymphocytes (PBLs) from 10 head-injured patients and 10 matched controls were evaluated following in vitro stimulation with the T-cell mitogen, phytohemagglutinin (PHA). Subsets of helper T cells evaluated included activated helper (CD4+/CD25+) T cells; helper/inducer (CD4+/CDw29+) T cells, which enhance cellular immune activity; and suppressor/inducer (CD4+/CD45R+) T-cells, which induce suppressor (CD8+) T-cells. In addition, the effect of intraventricular fluid (IVF) on PHA-stimulated in vitro CD4 and CD25 expression was investigated to determine whether severe head injury results in the production of mediators within the central nervous system capable of affecting T-cell activation. The results of this study indicate that isolated severe head injury selectively reduces the ability of PHA-stimulated PBLs to express the helper/inducer (CD4+/CDw29+) T-cell (p = 0.023) and activated helper (CD4+/CD25+) T-cell (P = 0.041) phenotypes. There was no significant change in PHA-stimulated CD4 or CD25 expression following incubation of PBLs with intraventricular fluid (IVF) from head-injured patients. The relationship between these changes in specific helper T-cell subpopulations and the infectious complications of severe head injury are discussed.     

Th1/Th2 balance and CD45-positive T cell subsets in primary nephrotic syndrome

T cells are involved in the pathogenesis of nephrotic syndrome (NS). The aim of the study was to determine whether the activity of T-helper-1 (Th1) and T-helper-2 (Th2) cells and the distribution of the lymphocyte subsets, namely CD45RA+CD4+ (”naive” helper T cells, suppressor-inducer), CD45RA+CD8+ (”naive” suppressor T cells, suppressor-effector), CD45RO+CD4+ (”memory” helper T cells), are predictive for steroid sensitivity in children with primary NS. These parameters were assessed at the onset of disease, before initiation of steroid therapy. Two groups of NS children were retrospectively formed according to steroid sensitivity (SS) or resistance (SR). The activity of Th1 and Th2 cells was defined by the production of interleukin-2 (IL-2), interferon-γ, IL-4, and IL-10 in the supernatants of CD4+ T cell cultures activated with autologous monocytes presenting tetanus toxoid (TT). Peripheral lymphocyte subsets were determined using double- or triple-color flow cytometry. In SS children with NS we found a decreased proliferative response of CD4+ T cells to TT stimulation, cytokine synthesis indicating the predominance of Th2 activity, and an increased percentage of activated suppressor-inducer (CD45RA+ CD4+CD25+, 5.18±0.8, P<0.001) and suppressor-effector (CD45RA+CD8+CD25+, 2.05±0.6, P<0.01) cells, with the concomitant reduction of activated memory cells (CD45RO+CD4+CD25+, 0.2±0.1, P<0.001). In children with SRNS we found an increased proliferative response of CD4+ T cells to TT, a rise in activated memory (CD45RO+CD4+CD25+, 3.82±0.7, P<0.01) and suppressor-inducer peripheral T cells (CD45RA+ CD4+CD25+, 3.85±0.6, P<0.01), but a low percentage of activated suppressor-effector (CD45RA+CD8+ CD25+, 0.5±0.2, P<0.05) T cells. We conclude that prior to treatment the distribution of lymphocyte subpopulations in peripheral blood together with Th1 and Th2 cell activity provides a useful tool for evaluating the likelihood of steroid sensitivity in patients with primary NS. Received: 3 November 1998 / Revised: 1 September 1999 / Accepted: 8 September 1999     

Allospecific CD154+ T Cells Associate with Rejection Risk After Pediatric Liver Transplantation

Antigen-specific T cells, which express CD154 rapidly, but remain untested in alloimmunity, were measured with flow cytometry in 16-h MLR of 58 identically-immunosuppressed children with liver transplantation (LTx), to identify Rejectors (who had experienced biopsy-proven rejection within 60 days posttransplantation). Thirty-one children were sampled once, cross-sectionally. Twenty-seven children were sampled longitudinally, pre-LTx, and at 1–60 and 61–200 days after LTx. Results were correlated with proliferative alloresponses measured by CFSE-dye dilution (n = 23), and CTLA4, a negative T-cell costimulator, which antagonizes CD154-mediated effects (n = 31). In cross-sectional observations, logistic regression and leave-one-out cross-validation identified donor-specific, CD154 + T-cytotoxic (Tc)-memory cells as best associated with rejection outcomes. In the longitudinal cohort, (1) the association between CD154 + Tc-memory cells and rejection outcomes was replicated with sensitivity/specificity 92.3%/84.6% for observations at 1–60 days, and (2) elevated pre-LTx CD154 + Tc-memory cell responses were associated with significantly increased incidence (p = 0.02) and hazard (HR = 7.355) of rejection in survival/proportional hazard analysis. CD154 expression correlated with proliferative alloresponses (r = 0.835, p = 7.1e-07), and inversely with CTLA4 expression of allospecific CD154 + Tc-memory cells (r =−0.706, p = 3.0e-05). Allospecific CD154 + T-helper-memory cells, not CD154 + Tc-memory, were inhibited by increasing Tacrolimus concentrations (p = 0.026). Collectively, allospecific CD154 + T cells provide an estimate of rejection risk in children with LTx.     

Correlation of in vitro CD4+ T helper cell function with clinical graft status in immunosuppressed kidney transplant recipients.

We recently identified three distinct T helper pathways which contribute to interleukin-2 (IL-2) production by human peripheral blood lymphocytes following stimulation with HLA alloantigens. In two of these pathways, CD4+ T helper cells respond to alloantigen using either self antigen-presenting cells (sAPC)* or allogeneic antigen-presenting cells (aAPC). A third pathway involves CD8+ T helper cells using aAPC. Previous in vitro studies have shown that the T helper pathway dependent on CD4+ T helper cells and sAPC (CD4-sAPC) is the most susceptible to suppression by cyclosporine. In the present study, we measured alloantigen-stimulated IL-2 production by PBL from 42 kidney transplant recipients to characterize the strength of the three T helper-APC pathways. In 58% of patients, a loss of the CD4-sAPC pathway was identified and was correlated with cyclosporine treatment. However, several patients not receiving cyclosporine also exhibited a similar loss of T helper cell function, suggesting that cyclosporine is not the only factor involved. Of 27 patients exhibiting depressed CD4-sAPC function, none had evidence of ongoing/recent graft rejection. In contrast, of 11 patients with no defects in the three pathways of in vitro T helper cell function, 6 had evidence of chronic graft rejection. Of considerable interest are the data obtained from a separate group of 4 patients who had episodes of acute rejection during the study. In each case, at the time of the rejection episode, all exhibited an intact CD4-sAPC pathway. However, samples tested prior to the rejection episode or after successful treatment of the rejection episode showed a depressed CD4-sAPC pathway. These results suggest that depression of the CD4-sAPC pathway represents adequate immunosuppression for graft retention and that patients not exhibiting such suppression are at increased risk for both acute and chronic graft rejection. These data may have relevance for diagnosis and/or prediction of graft rejection and may provide an in vitro method of monitoring the functional degree of immunosuppression in transplant recipients.     

Prognostic significance of T cell associated surface antigen density changes during OKT3 therapy of renal allograft rejection

We describe 12 acute rejection episodes in 11 cadaver donor renal allograft recipients who required OKT3* rescue treatment for steroid-resistant acute rejection (9) or for severe vascular (antibody-mediated) rejection (3). There were 3 treatment failures with subsequent graft loss. Using 2-color flow cytometry the total T (CD3), B (DR+), activated T (CD3DR), T helper/inducer (CD4), T cytotoxic/suppressor (CD8) and activated T cytotoxic cell (CD8DR) subsets were analyzed before, in mid course (5 to 7 days) and at the end of 12 to 14 days of therapy with 5 mg. OKT3 intravenously daily. In parallel changes in the density of such T cell associated antigens were analyzed. Significant decreases in the mean levels of the CD3 (p less than 0.001), CD3DR (p less than 0.05), CD4 (p less than 0.05), CD8 (p less than 0.05) and CD8DR (p less than 0.05) subsets were observed at mid course. A significant decrease in the density of CD3 was observed (p less than 0.0001). The surface antigen density of CD3DR, CD4 and CD8 had decreased by 160% (p less than 0.002), 383% (p less than 0.001) and 260% (p less than 0.001), respectively. At the end of treatment CD3 and CD4 subset levels increased by 425% and 240% (p less than 0.001 and p less than 0.005), respectively. In contrast, the CD3DR and CD8DR subset levels continued to decrease (p less than 0.05). A higher pre-treatment level of CD3DR and a less sharp decrease in CD3, CD4 and CD8 subsets were associated with a higher risk of treatment failure (p less than 0.05, p less than 0.01, p less than 0.05 and p less than 0.05, respectively). The mean decrease in the density of the CD3 marker in the lost grafts was significantly smaller compared to successful outcomes (p less than 0.001). The results of this preliminary study suggest that OKT3 affects T cell associated antigens other than CD3. Such may provide a sensitive prognostic index for the effectiveness of OKT3 therapy, and permit the identification of those patients who might require higher doses and/or duration of OKT3 therapy to enhance renal allograft salvage rates.     

Stimulation of distinct T cell subsets in MLR using human macrophage hybridomas differentially expressing class II antigens.

Recognition of class II antigens by alloreactive T cells is thought to be the major mechanism by which tissues undergo rejection. However, the specific role of the various class II antigens in the stimulation of these alloreactive cells remains to be elucidated. We have recently generated a series of human monocyte hybridomas that express distinct patterns of class II antigen expression. HLA-DR+ as well as HLA-DR-DP+DQ+ hybrids were capable of promoting T cell proliferation in a unidirectional mixed lymphocyte reaction. T cells stimulated by the HLA-DR+ clone 16.1 were predominantly of the CD4 (helper/inducer) phenotype. In contrast, T cells stimulated by the HLA-DR-DP+DQ+ clone 13 appear to reside in the CD8+ T cell subpopulation. Functional assessment of the T cell blasts generated in these cultures demonstrated a predominant helper T cell effect by those T cells stimulated by the HLA-DR+ clone 16.1, while suppressor cell activity was exhibited by T cells stimulated with the HLA-DR-DP+DQ+ clone 13. These data suggest that there may be a differential role for distinct class II molecules in the stimulation of T cell subpopulations.     

Decreased circulating CD4+CD25highFoxp3+ T cells during acute rejection in liver transplant patients

Background

CD4⁺CD25^highFoxP3⁺ regulatory T (Treg) cells have been implicated to maintain immunologic tolerance. They have been investigated in acute renal allografts rejection episodes (ARE). This study was performed to examine the frequency of peripheral blood (PB) CD4⁺CD25^highFoxP3⁺ Treg cells among liver transplantation patients with prior benign end-stage liver diseases in relation to ARE.

Methods

This prospective analysis of 55 patients who underwent liver transplantation from 2004 to 2009 did not include prisoners either as donors or recipients. PB was obtained from liver transplant patients longitudinally: pretransplantation, posttransplantation within 1 year, and at the time of an episode of ARE to measure by flow cytometry circulating CD4⁺CD25^highFoxP3⁺ T cells. Blood samples were drawn during ARE with concomitant liver biopsies. The rejector group was defined in the 14/55 cases who suffered an ARE; in the other patients with stable liver function were classified as the nonrejector group. We compared the number of circulating CD4⁺CD25^highFoxP3⁺ T cells between the 2 groups.

Results

There was no difference in the levels of circulating CD4⁺CD25^highFoxP3⁺ T cells pretransplantation. Interestingly, circulating CD4⁺CD25^highFoxP3⁺ T cells were significantly lower among the rejector compared with the nonrejector (2.23 ± 0.54% vs 2.99 ± 0.86%; P < .01). Longitudinal analysis revealed circulating CD4⁺CD25^highFoxP3⁺ T cells of patients in the rejector group to be significantly lower during rejection than during quiescence (2.23 ± 0.54% vs 3.68 ± 0.70%; P < .0001). The frequency of circulating CD4⁺CD25^highFoxP3⁺ T cells negatively correlated with a Rejection Activity Index (r = −0.80; P < .01).

Conclusion

Monitoring peripheral CD4⁺CD25^highFoxP3⁺ T cell levels may be useful to evaluate the immune state, potentially acting as a sensitive marker for ARE diagnosis among liver transplantation patients. Moreover, they may contribute to the mechanisms of Treg-mediated acceptance of liver transplantations.     

Usefulness of T-cell phenotype characterization in endomyocardial biopsy fragments from human cardiac allografts.

The mean numbers of cytotoxic/suppressor (CD8+) and helper/inducer (CD4+) T cells were determined in 111 successive endomyocardial biopsy fragments from eight cardiac allograft patients in an attempt to define their significance in the rejection process. Endomyocardial fragments from autopsy or donor hearts without myocarditis were evaluated as controls. The mean numbers of CD8+ and CD4+ T cells in the control group were 0.8 and 0.5 cells/field at x400 magnification, respectively. The mean numbers of CD8+ T cells per field in the cardiac allograft biopsies were 2.4, no rejection group; 5.4 mild rejection group; 11.1, moderate rejection group; and 4.9, resolving rejection group. The mean numbers of CD4+ T cells per field for the same groups were slightly lower than those of the CD8+ T cells. The number of CD8+ T cells per field reliably indicated the severity of rejection. Patients with normal numbers of CD8+ T cells and no evidence of rejection had better long-term outcomes (two or fewer moderate rejection episodes) than those with higher numbers. Analysis of the data suggests that the presence of two or fewer CD8+ T cells/field may be considered normal in the myocardial interstitium. The diagnosis of no evidence of rejection should be coupled to the presence of a normal number of CD8+ T cells. High numbers (greater than 10) of CD8+ T cells, even in absence of myocytolysis, should be treated more assertively, including the use of high doses of prednisone, because all our cases with high numbers showed a worse histologic picture at the subsequent biopsy.(ABSTRACT TRUNCATED AT 250 WORDS)     

Rejection of skin grafts and generation of cytotoxic T cells by mice depleted of L3T4+ cells

In mice, "helper/inducer" T cells can be depleted by treatment with a rat monoclonal antibody to the cell surface antigen, L3T4, which is homologous to the human antigen T4 (CD4). In order to examine the contribution of "helper/inducer" T cells to cellular immunity, C57BL/6 (H-2b) mice were treated weekly with 1 mg i.v. of a monoclonal antibody to L3T4. Three days after the first injection, the mice received skin grafts from BALB/c (H-2d) mice. The mice were then examined for skin graft rejection and for the development of cytotoxic cells. Treatment with anti-L3T4 prolonged skin graft survival from 9 to 18 days. Graft rejection was associated with the development of cellular cytotoxicity against H-2d targets. Cytotoxicity developed despite greater than or equal to 90% depletion of splenic L3T4+ cells. Allospecific cytotoxic T cells could also be generated in vitro from C57BL/6 spleen cells depleted of L3T4+ cells, when these were exposed in a mixed leukocyte culture to irradiated, T-cell-depleted, BALB/c spleen cells. In a mixed leukocyte culture using responder spleen cells from untreated C57BL/6 mice, both proliferation and interleukin 2 production were inhibited in the presence of antibody to L3T4 and, to a lesser extent, by antibody to Lyt-2. Complete inhibition was achieved by the presence of both antibodies. In a mixed leukocyte culture using responder spleen cells from C57BL/6 mice that had been treated with anti-L3T4, both proliferation and interleukin 2 production were inhibited largely by antibody to Lyt-2, although the presence of both anti-Lyt-2 and anti-L3T4 was most inhibitory. These findings indicate that graft rejection and cellular cytotoxicity can be generated in mice depleted of L3T4+ cells by methods that have previously been shown to abrogate humoral immunity. Cellular immunity appears to require few, if any, L3T4+ cells. These findings have implications for the clinical use of antibodies to "helper/inducer" T cells.     

Pretransplant Interferon‐γ Secretion by CMV‐Specific CD8+ T Cells Informs the Risk of CMV Replication After Transplantation

In this prospective study we analyzed pretransplant interferon‐γ secretion by cytomegalovirus (CMV)‐specific CD8+ T cells to assess its possible utility in determining the risk of CMV replication after solid organ transplantation. A total of 113 lung and kidney transplant patients were enrolled in the study but only 55 were evaluable. All CMV‐seronegative recipients were pretransplant “nonreactive” (IFNγ <0.2 IU/mL) (11/11), whereas 30/44 (68.2%) CMV‐seropositive (R+) recipients were “reactive” (IFNγ ≥0.2 IU/mL) and 14/44 (31.8%) were “nonreactive”. In the R(+) “nonreactive” group, 7/14 (50%) developed posttransplant CMV replication, whereas the virus replicated only in 4/30 (13.3%) of the R(+) “reactive” patients (p = 0.021). According to the best multivariate model, pretransplant “nonreactive” recipients receiving an organ from a CMV‐seropositive donor had a 10‐fold increased risk of CMV replication compared to pretransplant “reactive” recipients (adjusted OR 10.49, 95% CI 1.88–58.46). This model displayed good discrimination ability (AUC 0.80) and calibration (Hosmer–Lemeshow test, p = 0.92). Negative and positive predictive values were 83.7% and 75%, respectively. The accuracy of the model was 82%. Therefore, assessment of interferon‐γ secretion by cytomegalovirus (CMV)‐specific CD8+ T cells prior to transplantation is useful in informing the risk of posttransplant CMV replication in solid organ transplant patients.     

Monoclonal analysis of fine-needle aspiration biopsy in kidney allografts

To evaluate changes in T-lymphocyte subsets and DR expression on tubular cells, 74 fine-needle allograft aspirates (FNAB) were evaluated in 31 patients with cadaver kidney transplants. Monoclonal antibodies against T helper CD4+, cytotoxic/suppressor CD8+, and HLA-DR were used with an indirect alkaline-phosphatase-staining technique. Cases with acute rejection (n = 11) showed a significant increase of CD8+: CD4+ ratio versus those with stable function (n = 21), acute tubular necrosis (n = 10) or CsA toxicity (n = 7) (ANOVA F = 10; P less than 0.01). Cases with chronic rejection or CMV infection showed no differences in the CD8+: CD4+ ratio with the other groups. DR expression on tubular cells was frequently found in cases of acute rejection, chronic rejection and CMV (73%, 66%, and 43% respectively), occasionally found in CsA toxicity (14%), but never seen in controls or ATN. Both tests, the CD8+: CD4+ ratio and the DR expression on tubular cells, had a high sensitivity and specificity in differentiating acute rejection versus controls, acute tubular necrosis, and CsA toxicity. When both tests are taken together no case without rejection showed a CD8+:CD4+ ratio greater than 1.6 and DR expression on tubular cells. Cases with acute rejection who lost the graft (n = 6) had a CD8+:CD4+ ratio significantly greater than those who responded to antirejection therapy (n = 5) (t = 2.9; P less than 0.05).     

乳腺癌患者外周血CD4+CD25+Foxp3+调节性T细胞水平的检测及意义

目的：探讨乳腺癌患者外周血CD4+CD25+Foxp3+调节性T细胞（Treg）水平检测的意义。 方法：流式细胞术检测74例乳腺癌患者与30例健康对照者外周血CD4+CD25+Foxp3+Treg占CD4+T细胞百分比,分析CD4+CD25+Foxp3+Treg细胞水平与乳腺癌患者临床病理特征及相关免疫组化指标的关系。 结果：乳腺癌患者外周血CD4+CD25+Foxp3+Treg占CD4+T细胞的百分比高于健康对照者[（9.15± 2.24）% vs.（2.29±1.36）%],差异有统计学意义（P<0.05）。统计分析显示,乳腺癌患者外周血CD4+CD25+Foxp3+Treg细胞水平与肿瘤组织学分级、淋巴结转移、pTNM分期以及HER-2、pS2、nm23的表达有关（均P<0.05）,而与肿瘤大小、病理类型以及雌激素受体（ER）、孕激素受体（PR）、p53、Ki-67表达无关（均P>0.05）。进一步相关性分析显示,CD4+CD25+Foxp3+Treg细胞水平与肿瘤组织学分级、淋巴结转移数、pTNM分期、HER-2的表达呈正相关（r=0.583,r=0.333,r=0.919,r=0.604,均P<0.05）而与pS2、nm23表达呈负相关（r=-0.229,r=-0.401,均P<0.05）。 结论：乳腺癌患者外周血CD4+CD25+Foxp3+Treg细胞水平升高,并与与乳腺癌的进展、转移密切相关,对其检测可能有助于患者预后及治疗效果的评估。

     

Long‐term survival of pig‐to‐rhesus macaque renal xenografts is dependent on CD4 T cell depletion

The shortage of available organs remains the greatest barrier to expanding access to transplant. Despite advances in genetic editing and immunosuppression, survival in experimental models of kidney xenotransplant has generally been limited to <100 days. We found that pretransplant selection of recipients with low titers of anti‐pig antibodies significantly improved survival in a pig‐to–rhesus macaque kidney transplant model (6 days vs median survival time 235 days). Immunosuppression included transient pan–T cell depletion and an anti‐CD154–based maintenance regimen. Selective depletion of CD4⁺ T cells but not CD8⁺ T cells resulted in long‐term survival (median survival time >400 days vs 6 days). These studies suggested that CD4⁺ T cells may have a more prominent role in xenograft rejection compared with CD8⁺ T cells. Although animals that received selective depletion of CD8⁺ T cells showed signs of early cellular rejection (marked CD4⁺ infiltrates), animals receiving selective CD4⁺ depletion exhibited normal biopsy results until late, when signs of chronic antibody rejection were present. In vitro study results suggested that rhesus CD4⁺ T cells required the presence of SLA class II to mount an effective proliferative response. The combination of low pretransplant anti‐pig antibody and CD4 depletion resulted in consistent, long‐term xenograft survival.