Ajda T. Rowshani

Emergence of a CD4+CD28− Granzyme B+, Cytomegalovirus-Specific T Cell Subset after Recovery of Primary Cytomegalovirus Infection

Cytotoxic CD4+CD28− T cells form a rare subset in human peripheral blood. The presence of CD4+CD28− cells has been associated with chronic viral infections, but how these particular cells are generated is unknown. In this study, we show that in primary CMV infections, CD4+CD28− T cells emerge just after cessation of the viral load, indicating that infection with CMV triggers the formation of CD4+CD28− T cells. In line with this, we found these cells only in CMV-infected persons. CD4+CD28− cells had an Ag-primed phenotype and expressed the cytolytic molecules granzyme B and perforin. Importantly, CD4+CD28− cells were to a large extent CMV-specific because proliferation was only induced by CMV-Ag, but not by recall Ags such as purified protein derivative or tetanus toxoid. CD4+CD28− cells only produced IFN-γ after stimulation with CMV-Ag, whereas CD4+CD28+ cells also produced IFN-γ in response to varicella-zoster virus and purified protein derivative. Thus, CD4+CD28− T cells emerge as a consequence of CMV infection.

Clinical and immunologic aspects of cytomegalovirus infection in solid organ transplant recipients.

Primary cytomegalovirus (CMV) infection is a major cause of morbidity and mortality in recipients after solid organ transplantation (SOT). Widespread and prolonged use of antiviral drugs has changed the natural course of CMV disease by delaying its onset and causing drug resistance. CMV induces a strong cellular immune response, even in immunosuppressed patients, and has developed strategies to evade this immune surveillance. This review summarizes challenges in managing CMV infection in transplant recipients and highlights current insights in the cellular immune response against CMV.

CD103 is a marker for alloantigen-induced regulatory CD8(+) T cells

The αEβ7 integrin CD103 may direct lymphocytes to its ligand E-cadherin. CD103 is expressed on T cells in lung and gut and on allograft-infiltrating T cells. Moreover, recent studies have documented expression of CD103 on CD4+ regulatory T cells. Approximately 4% of circulating CD8+ T cells bear the CD103 molecule. In this study, we show that the absence or presence of CD103 was a stable trait when purified CD103− and CD103+CD8+ T cell subsets were stimulated with a combination of CD3 and CD28 mAbs. In contrast, allostimulation induced CD103 expression on ∼25% of purified CD103−CD8+ T cells. Expression of CD103 on alloreactive cells was found to be augmented by IL-4, IL-10, or TGF-β and decreased by addition of IL-12 to MLCs. The alloantigen-induced CD103+CD8+ T cell population appeared to be polyclonal and retained CD103 expression after restimulation. Markedly, in vitro-expanded CD103+CD8+ T cells had low proliferative and cytotoxic capacity, yet produced considerable amounts of IL-10. Strikingly, they potently suppressed T cell proliferation in MLC via a cell-cell contact-dependent mechanism. Thus, human alloantigen-induced CD103+CD8+ T cells possess functional features of regulatory T cells.

Serine proteases of the human immune system in health and disease.

Serine proteases form a large family of protein-cleaving enzymes that play an essential role in processes like blood coagulation, apoptosis and inflammation. Immune cells express a wide variety of serine proteases such as granzymes in cytotoxic lymphocytes, neutrophil elastase, cathepsin G and proteinase 3 in neutrophils and chymase and tryptase in mast cells. Regulation of proteolysis induced by these serine proteases is essential to prevent self-induced damage. Hence, there are specialized serine protease inhibitors, serpins, which are broadly distributed. Here, we discuss the function of human serine proteases in inflammation, apoptosis and tissue remodeling. Furthermore, we address their impact on development and progression of immune mediated-diseases. Understanding the mode of action of serine proteases will help to unravel molecular processes involved in immunological disorders and will facilitate the identification of new therapeutic targets.

No Difference in Degree of Interstitial Sirius Red–Stained Area in Serial Biopsies from Area under Concentration-over-Time Curves–Guided Cyclosporine versus Tacrolimus-Treated Renal Transplant Recipients at One Year

Interstitial fibrosis is the main characteristic of chronic allograft nephropathy and long-term graft failure. Cyclosporin (CsA) is thought to be more fibrogenic than tacrolimus. In a prospective, randomized, multicenter trial using a calcineurin-sparing regimen, renal interstitial volume was compared in CsA- and tacrolimus-treated renal transplant recipients by image analysis of Sirius red (SR)-stained cortical areas in protocol biopsies obtained at 6 mo (n = 94) and 12 mo (n = 97) after transplantation. Immunosuppression consisted of CsA or tacrolimus, CD25 mAb, mycophenolate mofetil, and prednisolone. CsA therapy increased the 6-mo risk for subclinical rejection. The prevalence of subclinical rejection was 38.8% in the CsA-treated and 15.2% in the tacrolimus-treated patient group (P = 0.012). Strikingly, no difference in the degree of interstitial SR-stained area was detectable between the two treatment groups. In particular, previous subclinical rejection episodes did not influence the degree of interstitial volume. Also, no difference in GFR occurred at 1 yr, when the mean GFR mounted 63 ml/min. No significant differences in the degree of interstitial SR-stained area could be observed at 6 and 12 mo between CsA- and tacrolimus-treated renal transplant recipients. Although CsA-treated patients developed significantly more subclinical rejections at 6 mo, this did not influence the degree of SR staining or the change in renal function at 1 yr.

Advancement of mesenchymal stem cell therapy in solid organ transplantation (MISOT).

There is evolving interest in the use of mesenchymal stem cells (MSC) in solid organ transplantation. Pre-clinical transplantation models show efficacy of MSC in prolonging graft survival and a number of clinical studies are planned or underway. At a recent meeting of the MISOT consortium (MSC In Solid Organ Transplantation) the advances of these studies were evaluated and mechanisms underlying the potential effects of MSC discussed. Continued discussion is required for definition of safety and eventually efficacy endpoints for MSC therapy in solid organ transplantation.

Molecular Comparison of Calcineurin Inhibitor-Induced Fibrogenic Responses in Protocol Renal Transplant Biopsies

The calcineurin inhibitor cyclosporine (CsA) induces a fibrogenic response that may lead to scarring of the renal allograft. This study investigated whether tacrolimus, a novel calcineurin inhibitor, exerts fibrogenic effects to a similar extent. Sixty patients were enrolled in a randomized study: 29 received CsA, and 31 received tacrolimus. Patients were subjected to tailored exposure-controlled calcineurin inhibitor regimens. Protocol biopsies were obtained at the time of transplantation and 6 and 12 mo after transplantation. Cortical TGF-beta and collagens alpha1(I) and alpha1(III) mRNA steady-state levels were determined with real-time PCR. The extent of protein deposition of TGF-beta, alpha-smooth muscle actin, and interstitial collagens in the renal cortex was quantified with computer-assisted image analysis. The extent of interstitial collagen deposition measured with Sirius red and the accumulation of alpha-smooth muscle actin and TGF-beta protein after 6 and 12 mo were similar for both immunosuppressive regimens. mRNA levels of TGF-beta and collagens alpha1(I) and alpha1(III) were not significantly different in the treatment groups either. It is concluded that the fibrogenic response in renal allografts is similar in patients who receive CsA-based regimens and patients who receive tacrolimus-based regimens.

Untreated Rejection in 6-Month Protocol Biopsies Is Not Associated with Fibrosis in Serial Biopsies or with Loss of Graft Function

Donor age, calcineurin inhibitor nephrotoxicity, and acute rejection are the most significant predictors of chronic allograft nephropathy. Protocol biopsies, both in deceased- and living-donor renal grafts, have shown that cortical tubulointerstitial fibrosis correlates with graft survival and function. The impact of not treating subclinical acute rejection (SAR) is less clear. In this study, 126 de novo renal transplant recipients were randomly assigned to receive area-under-the-curve-controlled exposure of either a cyclosporine or a tacrolimus-based immunosuppressive regimen that included steroids, mycophenolate mofetil, and basiliximab induction. Protocol biopsies were taken before and 6 and 12 mo after transplantation. The prevalence of SAR was determined retrospectively. Fibrosis was evaluated by quantitative digital analysis of Sirius red staining in serial biopsies. Donor age correlated significantly with tubulointerstitial fibrosis in pretransplantation biopsies and inferior graft function at month 6 (rtau = -0.26; P = 0.033). Acute rejection incidence was 11.5%, and no clinical late rejection occurred. The prevalence of SAR at 6 mo was 30.8% but was not associated with differences in serial quantitative Sirius red staining at 6 or 12 mo, proteinuria, or progressive loss of GFR up to 2 yr. No differences were found in donor variables, histocompatibility, rejection history, or exposure of immunosuppressants. Controlled individualized calcineurin inhibitor exposure and subsequent tapering resulted in a low early acute rejection rate and prevented late acute rejection. Because, by design, we did not treat SAR, these results provide evidence that asymptomatic infiltrates in 6-mo surveillance biopsies may not be deleterious in the intermediate term. There is need for reliable biomarkers to prove that not all cell infiltrates are equivalent or that infiltrates may change with time.

A Randomized Controlled Trial Comparing the Efficacy of Cyp3a5 Genotype-Based With Body-Weight-Based Tacrolimus Dosing After Living Donor Kidney Transplantation.

Patients expressing the cytochrome P450 (CYP) 3A5 gene require a higher tacrolimus dose to achieve therapeutic exposure compared with nonexpressers. This randomized‐controlled study investigated whether adaptation of the tacrolimus starting dose according to CYP3A5 genotype increases the proportion of kidney transplant recipients being within the target tacrolimus predose concentration range (10–15 ng/mL) at first steady‐state. Two hundred forty living‐donor, renal transplant recipients were assigned to either receive a standard, body‐weight‐based or a CYP3A5 genotype‐based tacrolimus starting dose. At day 3, no difference in the proportion of patients having a tacrolimus exposure within the target range was observed between the standard‐dose and genotype‐based groups: 37.4% versus 35.6%, respectively; p = 0.79. The proportion of patients with a subtherapeutic (i.e. <10 ng/mL) or a supratherapeutic (i.e. >15 ng/mL) Tac predose concentration in the two groups was also not significantly different. The incidence of acute rejection was comparable between both groups (p = 0.82). Pharmacogenetic adaptation of the tacrolimus starting dose does not increase the number of patients having therapeutic tacrolimus exposure early after transplantation and does not lead to improved clinical outcome in a low immunological risk population.

Human Cytomegalovirus Infection Induces a Rapid and Sustained Change in the Expression of NK Cell Receptors on CD8+ T Cells

The CD8+ T cell compartment of human CMV-seropositive individuals characteristically contains a high proportion of cells that express NK cell receptors (NKRs) which may contribute to the surveillance of virus-infected cells. To test whether this enhanced expression is a direct and immediate result of CMV infection, we used DNA microarrays to analyze putative changes in the RNA expression level of 39 NKRs in CMV-specific CD8+ T cells of renal transplant recipients experiencing primary CMV infection. Already in the acute phase of infection 29 NKRs were induced, of which 19 remained high 1 year after cessation of viral replication. Activating and inhibitory NKRs were induced to a similar extent. Detailed longitudinal flow cytometric analyses confirmed NKR changes at the protein level. Strikingly, a strong induction of CD94 on CD3+ T cells was observed with surface expression of activating CD94dim NKG2C dimers appearing before inhibitory CD94bright NKG2A ones. After the acute phase of infection, the balance between inhibitory and activating receptors did not change. Thus, CMV infection induces a rapid and lasting change in the expression of NKRs on human CD8+ T cells.