Decoding functional hematopoietic progenitor cells in the adult human lung.

The bone marrow is the main site of blood cell production in adults, however, rare pools of hematopoietic stem and progenitor cells with self-renewal and differentiation potential have been found in extramedullary organs. The lung is primarily known for its role in gas exchange but has recently been described as a site of blood production in mice. Here, we show that functional hematopoietic precursors reside in the extravascular spaces of the human lung, at a frequency similar to the bone marrow, and are capable of proliferation and engraftment. The organ-specific gene signature of pulmonary and medullary CD34+ hematopoietic progenitors indicates greater baseline activation of immune, megakaryocyte/platelet and erythroid-related pathways in lung progenitors. Spatial transcriptomics mapped blood progenitors in the lung to a vascular-rich alveolar interstitium niche. These results identify the lung as a pool for uniquely programmed blood stem and progenitor cells with the potential to support hematopoiesis in humans.

Sepsis promotes splenic production of a protective platelet pool with high CD40 ligand expression.

Platelets have a wide range of functions including critical roles in hemostasis, thrombosis, and immunity. We hypothesized that during acute inflammation, such as in life-threatening sepsis, there are fundamental changes in the sites of platelet production and phenotypes of resultant platelets. Here, we showed during sepsis that the spleen was a major site of megakaryopoiesis and platelet production. Sepsis provoked an adrenergic-dependent mobilization of megakaryocyte-erythrocyte progenitors (MEPs) from the bone marrow to the spleen, where IL-3 induced their differentiation into megakaryocytes (MKs). In the spleen, immune-skewed MKs produced a CD40 ligandhi platelet population with potent immunomodulatory functions. Transfusions of post-sepsis platelets enriched from splenic production enhanced immune responses and reduced overall mortality in sepsis-challenged animals. These findings identify a spleen-derived protective platelet population that may be broadly immunomodulatory in acute inflammatory states such as sepsis.

Clinical and Pathologic Feature of Patients With Early Versus Late Active Antibody-Mediated Rejection After Kidney Transplantation: A Single-Center Experience.

OBJECTIVE: Active antibody-mediated rejection (aABMR), particularly late aABMR, remains a major challenge for long-term renal allograft survival. This single-center retrospective study aimed to compare clinical features between early vs late aABMR and to identify risk factors for allograft failure among patients with aABMR. METHOD: Forty-one patients diagnosed with aABMR at our hospital were included and were divided into 2 groups: early aABMR (≤6 months; n = 10) vs late aABMR (>6 months; n = 31) based on the time from transplant to diagnosis. Their clinical and pathologic data were compared. This study was performed in compliance with the Helsinki Congress and the Declaration of Istanbul. RESULTS: Of 10 patients with early aABMR, none had allograft failure, whereas 8 of 31 patients with late aABMR had developed allograft failure at the time of follow-up (25.8%). At the time of biopsy, patients with early aABMR had higher positive grade in urine occult blood test than patients with late aABMR (P = .01); however, the late aABMR group displayed more intensive interstitial fibrosis and tubular atrophy (P = .03) and more frequent HLA-DQ-type donor-specific antibodies. Interestingly, donor-specific antibody conversion from positive to negative was not associated with C4d grade but was correlated with time from transplant to biopsy. Multivariate Cox regression analysis indicated that high levels of serum creatinine or proteinuria and concomitant T-cell-mediated rejection were independent risk factors for allograft failure in patients with aABMR. CONCLUSION: These data not only confirm that early aABMR has better clinical outcomes than late aABMR but highlight the importance of early diagnostic biopsy and early therapeutic interventions in ABMR, particularly in patients with high levels of serum creatinine or proteinuria in the early posttransplant phase.

A novel murine model of differentiation-mediated cytomegalovirus reactivation from latently infected bone marrow haematopoietic cells.

CD34+ myeloid lineage progenitor cells are an important reservoir of latent human cytomegalovirus (HCMV), and differentiation to macrophages or dendritic cells (DCs) is known to cause reactivation of latent virus. Due to its species-specificity, murine models have been used to study mouse CMV (MCMV) latency and reactivation in vivo. While previous studies have shown that MCMV genomic DNA can be detected in the bone marrow (BM) of latently infected mice, the identity of these cells has not been defined. Therefore, we sought to identify and enrich for cellular sites of MCMV latency in the BM haematopoietic system, and to explore the potential for establishing an in vitro model for reactivation of latent MCMV. We studied the kinetics and cellular characteristics of acute infection and establishment of latency in the BM of mice. We found that while MCMV can infect a broad range of haematopoietic BM cells (BMCs), latent virus is only detectable in haematopoietic stem cells (HSCs), myeloid progenitor cells, monocytes and DC-enriched cell subsets. Using three separate approaches, MCMV reactivation was detected in association with differentiation into DC-enriched BMCs cultured in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 4 (IL-4) followed by lipopolysaccharide (LPS) treatment. In summary, we have defined the kinetics and cellular profile of MCMV infection followed by the natural establishment of latency in vivo in the mouse BM haematopoietic system, including the haematopoietic phenotypes of cells that are permissive to acute infection, establish and harbour detectable latent virus, and can be stimulated to reactivate following DC enrichment and differentiation, followed by treatment with LPS.

A Mouse Model of Vascularized Heterotopic Spleen Transplantation for Studying Spleen Cell Biology and Transplant Immunity.

The spleen is a unique lymphoid organ that plays a critical role in the homeostasis of the immune and hematopoietic systems. Patients that have undergone splenectomy regardless of precipitating causes are prone to develop an overwhelming post-splenectomy infection and experience increased risks of deep venous thrombosis and malignancies. Recently, epidemiological studies indicated that splenectomy might be associated with the occurrence of cardiovascular diseases, suggesting that physiological functions of the spleen have not yet been fully recognized. Here, we introduce a mouse model of vascularized heterotopic spleen transplantation, which not only can be utilized to study the function and behavioral activity of splenic immune cell subsets in different biologic processes, but also can be a powerful tool to test the therapeutic potential of spleen transplantation in certain diseases. The main surgical steps of this model include donor spleen harvest, the removal of recipient native spleen, and spleen graft revascularization. Using congenic mouse strains (e.g., mice with CD45.1/CD45.2 backgrounds), we observed that after syngeneic transplantation, both donor-derived splenic lymphocytes and myeloid cells migrated out of the graft as early as post-operative day 1, concomitant with the influx of multiple types of recipient cells, thus generating a unique chimera.  Despite relatively challenging techniques, this procedure can be performed with >90% success rate. This model allows tracking the fate, longevity, and function of splenocytes during steady state and in a disease setting following a spleen transplantation, thereby offering a great opportunity to discover the distinct role for spleen-derived immune cells in different disease processes.

Evaluation of quality of kidneys from donation after circulatory death/expanded criteria donors by parameters of machine perfusion.

AIM: To investigate whether the parameters of machine perfusion could predict the quality of kidneys from donation after circulatory death (DCD) donors and expanded criteria donors (ECD). METHODS: Fifty-eight kidneys from DCD/ECD donors were harvested in our hospital from July 2011 to August 2014. All kidneys were preserved with machine perfusion (Life Port), and parameters of machine perfusion were collected. All kidneys were biopsied before transplantation. The primary endpoints were delayed graft function (DGF), graft loss and patient death. RESULTS: After kidney transplantation, 26 patients (44.8%) had DGF. We chose 1 h RI as a predictive parameter to predict DGF after transplant, and made the ROC curve. The ROC curve showed that 1 h RI = 0.4 was the best cut-off point for predicting DGF after transplant. The sensitivity was 61.54%, and the specificity was 81.25%. Fifty-eight recipients were divided into two groups according to 1 h RI of machine perfusion. 22 cases in high RI group (RI > 0.4) and 36 cases in low RI group (RI ≤0.4). DGF rate was significantly higher in the high RI group (72.7% vs. 27.8%). One year serum creatinine levels were also significantly higher in the high RI group (P < 0.05). Acute rejection rate and 1 year graft and patient survival were comparable. CONCLUSIONS: One hour RI of machine perfusion is associated with DGF and 1 year graft function in DCD/ECD kidney transplantation, and may be a non-invasive tool for evaluating quality of DCD/ECD kidneys.

The BTK Inhibitor Ibrutinib (PCI-32765) Overcomes Paclitaxel Resistance in ABCB1- and ABCC10-Overexpressing Cells and Tumors.

Paclitaxel is one of the most widely used antineoplastic drugs in the clinic. Unfortunately, the occurrence of cellular resistance has limited its efficacy and application. The ATP-binding cassette subfamily B member 1 (ABCB1/P-glycoprotein) and subfamily C member 10 (ABCC10/MRP7) are the major membrane protein transporters responsible for the efflux of paclitaxel, constituting one of the most important mechanisms of paclitaxel resistance. Here, we demonstrated that the Bruton tyrosine kinase inhibitor, ibrutinib, significantly enhanced the antitumor activity of paclitaxel by antagonizing the efflux function of ABCB1 and ABCC10 in cells overexpressing these transporters. Furthermore, we demonstrated that the ABCB1 or ABCC10 protein expression was not altered after treatment with ibrutinib for up to 72 hours using Western blot analysis. However, the ATPase activity of ABCB1 was significantly stimulated by treatment with ibrutinib. Molecular docking analysis suggested the binding conformation of ibrutinib within the large cavity of the transmembrane region of ABCB1. Importantly, ibrutinib could effectively enhance paclitaxel-induced inhibition on the growth of ABCB1- and ABCC10-overexpressing tumors in nude athymic mice. These results demonstrate that the combination of ibrutinib and paclitaxel can effectively antagonize ABCB1- or ABCC10-mediated paclitaxel resistance that could be of great clinical interest. Mol Cancer Ther; 16(6); 1021-30. ©2017 AACR.

Ethylene carbodiimide-fixed donor splenocytes combined with α-1 antitrypsin induce indefinite donor-specific protection to mice cardiac allografts.

Peritransplant infusion of ethylene carbodiimide-fixed donor splenocytes (ECDI-SPs) induces protection of islet and cardiac allografts. However, pro-inflammatory cytokine production during the peritransplantation period may negate the effect of ECDI-SPs. Therefore, we hypothesized that blocking pro-inflammatory cytokine secretion while increasing levels of anti-inflammatory cytokines would enhance the tolerance-induced efficacy of ECDI-SPs. The objective of this study was to determine the effectiveness of using ECDI-SPs combined with a short course of α1-antitrypsin (AAT) for induction of tolerance. Using a mice cardiac transplant model, we demonstrated that ECDI-SPs + AAT effectively induced indefinite mice cardiac allograft protection in a donor-specific fashion. This effect was accompanied by modulation of cytokines through decreasing levels of pro-inflammatory cytokines (including IFN-γ, TNF-α, IL-1ß, IL-6, IL-17, and IL-23) and increasing levels of anti-inflammatory cytokines (including IL-10, IL-13, and TGF-ß), and by inhibition of effector T cells (Teff) and expansion of regulatory T cells (Tregs). Therefore, we concluded that combined ECDI-SPs and AAT appeared to modulate the expression of cytokines and regulate the Teff:Treg balance to create a support milieu for graft protection. Our strategy of combining ECDI-SPs and AAT provides a promising approach for inducing donor-specific transplant tolerance.