Direct Tumor Killing and Immunotherapy through Anti-SerpinB9 Therapy.

Cancer therapies kill tumors either directly or indirectly by evoking immune responses and have been combined with varying levels of success. Here, we describe a paradigm to control cancer growth that is based on both direct tumor killing and the triggering of protective immunity. Genetic ablation of serine protease inhibitor SerpinB9 (Sb9) results in the death of tumor cells in a granzyme B (GrB)-dependent manner. Sb9-deficient mice exhibited protective T cell-based host immunity to tumors in association with a decline in GrB-expressing immunosuppressive cells within the tumor microenvironment (TME). Maximal protection against tumor development was observed when the tumor and host were deficient in Sb9. The therapeutic utility of Sb9 inhibition was demonstrated by the control of tumor growth, resulting in increased survival times in mice. Our studies describe a molecular target that permits a combination of tumor ablation, interference within the TME, and immunotherapy in one potential modality.

Lactate Dehydrogenase and Hemorrhagic Stroke During Extracorporeal Membrane Oxygenation for COVID-19.

PURPOSE: Hemorrhagic stroke (HS) is a devastating complication during extracorporeal membrane oxygenation (ECMO) but markers of risk stratification during COVID-19 are unknown. Lactate dehydrogenase (LDH) is a readily available biomarker of cell injury and permeability. We sought to determine whether an elevated LDH before ECMO placement is related to the occurrence of HS during ECMO for COVID-19. METHODS: Adult patients with COVID-19 requiring ECMO between March 2020 and February 2022 were included. LDH values prior to ECMO placement were captured. Patients were categorized into high (> 750 U/L) or low (≤ 750 U/L) LDH groups. Multivariable regression modeling was used to determine the association between LDH and HS during ECMO. RESULTS: There were 520 patients that underwent ECMO placement in 17 centers and 384 had an available LDH. Of whom, 122 (32%) had a high LDH. The overall incidence of HS was 10.9%, and patients with high LDH had a higher incidence of HS than those with low LDH level (17% vs 8%, p = 0.007). At 100 days, the probability of a HS was 40% in the high LDH group and 23% in those with a low LDH, p = 0.002. After adjustment for clinical covariates, high LDH remained associated with subsequent HS (aHR: 2.64, 95% CI 1.39-4.92). Findings were similar when restricting to patients supported by venovenous ECMO only. CONCLUSION: Elevated LDH prior to ECMO cannulation is associated with a higher incidence of HS during device support. LDH can risk stratify cases for impending cerebral bleeding during ECMO.

miR-21 antagonism reprograms macrophage metabolism and abrogates chronic allograft vasculopathy.

Despite much progress in improving graft outcome during cardiac transplantation, chronic allograft vasculopathy (CAV) remains an impediment to long-term graft survival. MicroRNAs (miRNAs) emerged as regulators of the immune response. Here, we aimed to examine the miRNA network involved in CAV. miRNA profiling of heart samples obtained from a murine model of CAV and from cardiac-transplanted patients with CAV demonstrated that miR-21 was most significantly expressed and was primarily localized to macrophages. Interestingly, macrophage depletion with clodronate did not significantly prolong allograft survival in mice, while conditional deletion of miR-21 in macrophages or the use of a specific miR-21 antagomir resulted in indefinite cardiac allograft survival and abrogated CAV. The immunophenotype, secretome, ability to phagocytose, migration, and antigen presentation of macrophages were unaffected by miR-21 targeting, while macrophage metabolism was reprogrammed, with a shift toward oxidative phosphorylation in naïve macrophages and with an inhibition of glycolysis in pro-inflammatory macrophages. The aforementioned effects resulted in an increase in M2-like macrophages, which could be reverted by the addition of L-arginine. RNA-seq analysis confirmed alterations in arginase-associated pathways associated with miR-21 antagonism. In conclusion, miR-21 is overexpressed in murine and human CAV, and its targeting delays CAV onset by reprogramming macrophages metabolism.

Urine podoplanin heralds the onset of ischemia-reperfusion injury of the kidney.

Ischemia-reperfusion injury represents one of the most common causes of acute kidney injury, a serious and often deadly condition that affects up to 20% of all hospitalized patients in the United States. However, the current standard assay used universally for the diagnosis of acute kidney injury, serum creatinine, does not detect renal damage early in its course. Serendipitously, we found that the immunofluorescent signal of the constitutive podocyte marker podoplanin fades in the glomerulus and intensifies in the tubulointerstitial compartment of the kidney shortly after ischemia-reperfusion injury in 8- to 10-wk-old male C57Bl/6j mice. Therefore, we sought to define the appearance and course of the podoplanin-positive signal in the kidney after ischemia-reperfusion injury. The tubulointerstitial podoplanin-positive signal increased as early as 2 h but persisted for 7 days after ischemia-reperfusion injury. In addition, the strength of this tubulointerstitial signal was directly proportional to the severity of ischemia, and its location shifted from the tubules to interstitial cells over time. Finally, we detected podoplanin in the urine of mice after ischemia, and we observed that an increase in the urine podoplanin-to-creatinine ratio correlated strongly with the onset of renal ischemia-reperfusion injury. Our findings indicate that the measurement of urine podoplanin harbors promising potential for use as a novel biomarker for the early detection of ischemia-reperfusion injury of the kidney.

Activation of fibroblastic reticular cells in kidney lymph node during crescentic glomerulonephritis.

Crescentic glomerulonephritis is an inflammatory condition characterized by rapid deterioration of kidney function. Previous studies of crescentic glomerulonephritis have focused on immune activation in the kidney. However, the role of fibroblastic reticular cells, which reside in the stromal compartment of the kidney lymph node, has not been studied in this condition. We investigated the activation of kidney lymph node-resident fibroblastic reticular cells in nephrotoxic serum nephritis, a classic murine model of crescentic glomerulonephritis. We found that increased deposition of extracellular matrix fibers by fibroblastic reticular cells in the kidney lymph node was associated with the propagation of high endothelial venules, specialized blood vessels through which lymphocytes enter the lymph node, as well as with expansion of the lymphatic vasculature. The kidney lymph node also contained an expanding population of pro-inflammatory T cells. Removal of the kidney lymph node, depletion of fibroblastic reticular cells, and treatment with anti-podoplanin antibody each resulted in reduction of kidney injury. Our findings suggest that modulating the activity of fibroblastic reticular cells may be a novel therapeutic approach in crescentic glomerulonephritis.

Brief treatment with a highly selective immunoproteasome inhibitor promotes long-term cardiac allograft acceptance in mice.

Constitutive proteasomes (c-20S) are ubiquitously expressed cellular proteases that degrade polyubiquitinated proteins and regulate cell functions. An isoform of proteasome, the immunoproteasome (i-20S), is highly expressed in human T cells, dendritic cells (DCs), and B cells, suggesting that it could be a potential target for inflammatory diseases, including those involving autoimmunity and alloimmunity. Here, we describe DPLG3, a rationally designed, noncovalent inhibitor of the immunoproteasome chymotryptic subunit ß5i that has thousands-fold selectivity over constitutive ß5c. DPLG3 suppressed cytokine release from blood mononuclear cells and the activation of DCs and T cells, diminished accumulation of effector T cells, promoted expression of exhaustion and coinhibitory markers on T cells, and synergized with CTLA4-Ig to promote long-term acceptance of cardiac allografts across a major histocompatibility barrier. These findings demonstrate the potential value of using brief posttransplant immunoproteasome inhibition to entrain a long-term response favorable to allograft survival as part of an immunomodulatory regimen that is neither broadly immunosuppressive nor toxic.

Salt Accelerates Allograft Rejection through Serum- and Glucocorticoid-Regulated Kinase-1-Dependent Inhibition of Regulatory T Cells.

A high-salt diet (HSD) in humans is linked to a number of complications, including hypertension and cardiovascular events. Whether a HSD affects the immune response in transplantation is unknown. Using a murine transplantation model, we investigated the effect of NaCl on the alloimmune response in vitro and in vivo. Incremental NaCl concentrations in vitro augmented T cell proliferation in the settings of both polyclonal and allospecific stimulation. Feeding a HSD to C57BL/6 wild-type recipients of bm12 allografts led to accelerated cardiac allograft rejection, despite similar mean BP and serum sodium levels in HSD and normal salt diet (NSD) groups. The accelerated rejection was associated with a reduction in the proportion of CD4(+)Foxp3(+) regulatory T cells (Tregs) and a significant decrease in Treg proliferation, leading to an increased ratio of antigen-experienced CD4(+) T cells to Tregs in mice recipients of a HSD compared with mice recipients of a NSD. Because serum- and glucocorticoid-regulated kinase-1 (SGK1) has been proposed as a potential target of salt in immune cells, we fed a HSD to CD4(Cre)SGK1(fl/fl) B6-transplanted recipients and observed abrogation of the deleterious effect of a HSD in the absence of SGK1 on CD4(+) cells. In summary, we show that NaCl negatively affects the regulatory balance of T cells in transplantation and precipitates rejection in an SGK1-dependent manner.

[Intraoperative evaluation for residual mitral valve regurgitation; usefulness of the retrograde cardioprotective beating test].

Intraoperative evaluation is important for successful mitral valve plasty (MVP). We performed a saline injection test and a retrograde cardioprotective beating test (RC-beating test) for intraoperative evaluation. The concept of the RC- beating test is evaluation of residual mitral valve regurgitation( MR) under cardiac beating. A 66-year-old man with severe MR underwent MVP. The P3 chorda was ruptured and we performed quadrangular resection. The saline injection test showed trivial regurgitation. We then performed the RC-beating test and it revealed severe leakage from the posterior commissure(PC). Since the PC had a sclerotic change, another quadrangular resection was performed. Moreover,as the anterior leaflet( A3) was slightly elongated, the region was resected in an obtuse-angled triangle shape and repaired by suturing the edges. The final RC-beating test showed no residual leakage. The RC-beating test is useful for detecting residual mitral valve leakage.

Donation After Circulatory Death Heart Transplant: Current State and Future Directions.

Orthotopic heart transplant is the gold standard therapeutic intervention for patients with end-stage heart failure. Conventionally, heart transplant has relied on donation after brain death for organ recovery. Donation after circulatory death (DCD) is the donation of the heart after confirming that circulatory function has irreversibly ceased. DCD-orthotopic heart transplant differs from donation after brain death-orthotopic heart transplant in ways that carry implications for widespread adoption, including differences in organ recovery, storage and ethical considerations surrounding normothermic regional perfusion with DCD. Despite these differences, DCD has shown promising early outcomes, augmenting the donor pool and allowing more individuals to benefit from orthotopic heart transplant. This review aims to present the current state and future trajectory of DCD-heart transplant, examine key differences between DCD and donation after brain death, including clinical experiences and innovations in methodologies, and address the ongoing ethical challenges surrounding the new frontier in heart transplant with DCD donors.

Use of Extracorporeal Membrane Oxygenation for Primary Graft Dysfunction After Cardiac Transplantation: Results of an A Priori Ventless Approach.

Primary graft dysfunction (PGD) after cardiac transplantation is a devastating complication with increasing frequency lately in the setting of donation after circulatory death (DCD). Severe PGD is commonly treated with extracorporeal membrane oxygenation (ECMO) using central or peripheral cannulation. We retrospectively reviewed the outcomes of PGD after cardiac transplantation requiring ECMO support at our center from 2015 to 2020, focused on our now preferential approach using peripheral cannulation without a priori venting. During the study period, 255 patients underwent heart transplantation at our center and 26 (10.2%) of them required ECMO for PGD. Of 24 patients cannulated peripherally 19 (79%) were alive at 30 days and 17 (71%) 1 year after transplant; two additional patients underwent central ECMO cannulation due to unfavorable size of femoral vessels and concern for limb ischemia. Successful decannulation with full graft function recovery occurred in 22 of 24 (92%) patients cannulated peripherally. Six of them had an indwelling intra-aortic balloon pump placed before the transplantation. None of the other 18 patients received a ventricular vent. In conclusion, the use of an a priori peripheral and ventless ECMO approach in patients with PGD after heart transplant is an effective strategy associated with high rates of graft recovery and survival.

Infradiaphragmatic malperfusion of acute aortic dissection associated with previous abdominal aortic aneurysm repair.

PURPOSE: To evaluate the association of previous abdominal aortic aneurysm (AAA) graft replacement with infradiaphragmatic malperfusion in patients with acute aortic dissection. METHODS: Between November 2006 and June 2011, 133 patients were referred to our hospital for management of acute aortic dissection. Eight (6.0 %) of these patients had undergone AAA graft replacement prior to the acute aortic dissection. We compared the computed tomography (CT) images of these 8 patients with those of the remaining 125 patients without previous AAA graft replacement, in terms of organ ischemia as a complication induced by acute aortic dissection. RESULTS: Infradiaphragmatic malperfusion from acute aortic dissection was confirmed in four of the eight patients who had undergone AAA graft replacement. Contrasted CT scan images indicated that the main cause of infradiaphragmatic malperfusion was collapse of the true lumen from compression by the false lumen into the suprarenal aorta. Although there was no significant difference between the groups in terms of cerebral ischemia and myocardial ischemia, bilateral leg ischemia and visceral ischemia occurred more frequently in the patients who had undergone AAA graft replacement. CONCLUSION: Previous AAA graft replacement is a risk factor for infradiaphragmatic malperfusion in patients with acute aortic dissection.

[Neuroleptic malignant syndrome after cardiac surgery].

A 64-year-old man without any psychiatric disease, including Parkinson's disease underwent aortic valve replacement and mitral valve replacement for rheumatic valvular disease. One day after the cardiac surgery, he developed hyperthermina, muscle rigidity, coma and delirium, and his serum creatine kinase (CK) level was elevated. In spite of his negative brain computed tomography(CT), his consciousness remained unclear. He had received diazepam, flunitrazepam and buprenorphine after the cardiac surgery because of his hyper-reactivity. Although these drugs were not typical antipsychotics' causing neuroleptic malignant syndrome (NMS), NMS was strongly suspected because of his clinical appearance. Dantrolene was administered in a dose of 60 mg per day and he recovered consciousness and his CK level began to decrease. We reported a case of neuroleptic malignant syndrome after cardiac surgery.

[Surgical treatment of coronary aneurysm after percutaneous coronary intervention].

A 61-year-old man with acute myocardial infarction underwent percutaneous coronary intervention with stent for the left main coronary artery (LMT) and the left anterior descending artery (LAD). Three months later, we recognized the LMT aneurysm complicated with possible thrombus formation, which developed in size during 6 months. In addition, the LAD stent showed significant in-stent stenosis. For the purpose of supplying blood flow to the distal of LAD, and avoiding myocardial infarction due to distal thrombosis possibly originated from LMT aneurysm, we decided to perform surgical operation. On preoperative examination, this patient had an obstruction of the right internal carotid artery. Although the direct repair of LMT aneurysm requires conventional approach with cardiopulmonary bypass, we applied off-pump coronary artery bypass grafting( OPCAB) considering the risk of cerebrovascular event. Consequently, OPCAB was performed in usual fashion [right internal thoracic artery (RITA) -LAD, left internal thoracic artery-left circumflex artery (LITA-LCX)] followed by the ligation of the proximal of LAD and LCX without cardiopulmonary bypass. The patient had a good operative course.

Characterization of CD41+ cells in the lymph node.

Lymph nodes (LNs) are the critical sites of immunity, and the stromal cells of LNs are crucial to their function. Our understanding of the stromal compartment of the LN has deepened recently with the characterization of nontraditional stromal cells. CD41 (integrin αIIb) is known to be expressed by platelets and hematolymphoid cells. We identified two distinct populations of CD41+Lyve1+ and CD41+Lyve1- cells in the LNs. CD41+Lyve1- cells appear in the LN mostly at the later stages of the lives of mice. We identified CD41+ cells in human LNs as well. We demonstrated that murine CD41+ cells express mesodermal markers, such as Sca-1, CD105 and CD29, but lack platelet markers. We did not observe the presence of platelets around the HEVs or within proximity to fibroblastic reticular cells of the LN. Examination of thoracic duct lymph fluid showed the presence of CD41+Lyve1- cells, suggesting that these cells recirculate throughout the body. FTY720 reduced their trafficking to lymph fluid, suggesting that their egress is controlled by the S1P1 pathway. CD41+Lyve1- cells of the LNs were sensitive to radiation, suggestive of their replicative nature. Single cell RNA sequencing data showed that the CD41+ cell population in naïve mouse LNs expressed largely stromal cell markers. Further studies are required to examine more deeply the role of CD41+ cells in the function of LNs.

ACTH treatment promotes murine cardiac allograft acceptance.

Heart transplantation is the optimal therapy for patients with end-stage heart disease, but its long-term outcome remains inadequate. Recent studies have highlighted the importance of the melanocortin receptors (MCRs) in inflammation, but how MCRs regulate the balance between alloreactive T cells and Tregs, and whether they impact chronic heart transplant rejection, is unknown. Here, we found that Tregs express MC2R, and MC2R expression was highest among all MCRs by Tregs. Our data indicate that adrenocorticotropic hormone (ACTH), the sole ligand for MC2R, promoted the formation of Tregs by increasing the expression of IL-2Rα (CD25) in CD4+ T cells and activation of STAT5 in CD4+CD25+ T cells. ACTH treatment also improved the survival of heart allografts and increased the formation of Tregs in CD28KO mice. ACTH treatment synergized with the tolerogenic effect of CTLA-4-Ig, resulting in long-term survival of heart allografts and an increase in intragraft Tregs. ACTH administration also demonstrated higher prolongation of heart allograft survival in transgenic mouse recipients with both complete KO and conditional KO of PI3Kγ in T cells. Finally, ACTH treatment reduced chronic rejection markedly. These data demonstrate that ACTH treatment improved heart transplant outcomes, and this effect correlated with an increase in Tregs.

Characterization of Leptin Receptor+ Stromal Cells in Lymph Node.

Lymph node (LN)-resident stromal cells play an essential role in the proper functioning of LNs. The stromal compartment of the LN undergoes significant compensatory changes to produce a milieu amenable for regulation of the immune response. We have identified a distinct population of leptin receptor-expressing (LepR+) stromal cells, located in the vicinity of the high endothelial venules (HEVs) and lymphatics. These LepR+ stromal cells expressed markers for fibroblastic reticular cells (FRCs), but they lacked markers for follicular dendritic cells (FDCs) and marginal reticular cells (MRCs). Leptin signaling deficiency led to heightened inflammatory responses within the LNs of db/db mice, leakiness of HEVs, and lymphatic fragmentation. Leptin signaling through the JAK/STAT pathway supported LN stromal cell survival and promoted the anti-inflammatory properties of these cells. Conditional knockout of the LepR+ stromal cells in LNs resulted in HEV and extracellular matrix (ECM) abnormalities. Treatment of ob/ob mice with an agonist leptin fusion protein restored the microarchitecture of LNs, reduced intra-LN inflammatory responses, and corrected metabolic abnormalities. Future studies are needed to study the importance of LN stomal cell dysfunction to the pathogenesis of inflammatory responses in type 2 diabetes (T2D) in humans.

[Idiopathic pulmonary trunk aneurysm].

Pulmonary trunk aneurysm is generally associated with congenital cardiac defects, pulmonary hypertension, or infection. Idiopathic pulmonary trunk aneurysm without any associated diseases is a rare lesion and has seldom been reported. Here, we report a case of a 68-year-old woman with idiopathic pulmonary trunk aneurysm. The maximum diameter of the aneurysm was 53 mm while she was 142 cm in height. We successfully performed aneurysmorrhaphy and her postoperative course was uneventful. Aneurysmorrhaphy was an effective technique for idiopathic pulmonary trunk aneurysm without pulmonary hypertention.

[Extirpation of a thrombus during pulmonary metastasectomy in a patient with chronic pulmonary thromboembolism].

A 57-year-old woman with chronic pulmonary thromboembolism was diagnosed to have metastatic lung tumors 6 months after an operation performed for colon cancer. There were no respiratory symptoms associated with the pulmonary thromboembolism. Computed tomography showed 2 pulmonary masses in the right middle and lower lobes, and a thrombus in the inferior trunk of the right pulmonary artery. Excision of the 2 pulmonary metastases was performed and the thrombus in the pulmonary artery was also removed at the same time. Partial improvement of the pulmonary blood flow was observed in the postoperative pulmonary scintigram. Chronic pulmonary thromboembolism without respiratory symptoms is not usually an indication for extirpation of thrombus. However, incidental extirpation of a pulmonary thrombus at the time of other pulmonary surgery should be considered, as it may yield an improvement of the pulmonary blood flow.

Cellular Mechanisms of Rejection of Optic and Sciatic Nerve Transplants: An Observational Study.

BACKGROUND: Organ transplantation is a standard therapeutic strategy for irreversible organ damage, but the utility of nerve transplantation remains generally unexplored, despite its potential benefit to a large patient population. Here, we aimed to establish a feasible preclinical mouse model for understanding the cellular mechanisms behind the rejection of peripheral and optic nerves. METHODS: We performed syngenic and allogenic transplantation of optic and sciatic nerves in mice by inserting the nerve grafts inside the kidney capsule, and we assessed the allografts for signs of rejection through 14 d following transplantation. Then, we assessed the efficacy of CTLA4 Ig, Rapamycin, and anti-CD3 antibody in suppressing immune cell infiltration of the nerve allografts. RESULTS: By 3 d posttransplantation, both sciatic and optic nerves transplanted from BALB/c mice into C57BL/6J recipients contained immune cell infiltrates, which included more CD11b+ macrophages than CD3+ T cells or B220+ B cells. Ex vivo immunogenicity assays demonstrated that sciatic nerves demonstrated higher alloreactivity in comparison with optic nerves. Interestingly, optic nerves contained higher populations of anti-inflammatory PD-L1+ cells than sciatic nerves. Treatment with anti-CD3 antibody reduced immune cell infiltrates in the optic nerve allograft, but exerted no significant effect in the sciatic nerve allograft. CONCLUSIONS: These findings establish the feasibility of a preclinical allogenic nerve transplantation model and provide the basis for future testing of directed, high-intensity immunosuppression in these mice.