Anomalous systemic and pulmonary veins—An unusual coexistence

A 12‐year‐old child presented with recurrent respiratory infections and was diagnosed with Scimitar syndrome. Drainage of IVC and course of aberrant arterial supply from aorta were not clear by echocardiogram, and hence, additional imaging was planned. CT unraveled the presence of a rare combination of anomalous pulmonary venous connection (APVC) to IVC, with near‐atresia/severe stenosis of IVC, superior to the drainage of scimitar vein. There was a prominent azygos vein with preferential contrast opacification on lower limb injection.     

Detection of cardiac myosin heavy chain-α-specific CD4 cells by using MHC class II/IA(k) tetramers in A/J mice

A/J mice bearing the H-2 allele IA(k) are highly susceptible to autoimmune myocarditis induced with cardiac myosin heavy chain (Myhc)-α 334-352, whereas B10.A mice carrying a similar allele IA(k) are relatively resistant, suggesting that the generation of Myhc-α-reactive T cell repertoires is influenced by genetic background. To enumerate the precursor frequencies of Myhc-α-specific CD4 T cells, we sought to create IA(k) tetramers for Myhc-α 334-352. Tetramers were created using approaches that involve covalent tethering of individual peptide sequences or exogenous loading of peptides into empty IA(k) molecules by peptide-exchange reaction. Using ribonuclease 43-56 tetramers as controls, we demonstrated that by flow cytometry (FC), Myhc-α 334-352 tetramers specifically bind myosin-reactive T cells. CD4 cells isolated from A/J mice immunized with Myhc-α 334-352 were used to optimize conditions for tetramer staining, and neuraminidase treatment prior to tetramer staining permitted the detection of Myhc-α-specific cells ex vivo. The reagents are useful tools for monitoring the frequency of Myhc-α-reactive CD4 cells and to determine their pathogenic potential at a single cell level by FC.     

Intricacies of cardiac damage in coxsackievirus B3 infection: Implications for therapy

Heart disease is the leading cause of death in humans, and myocarditis is one predominant cause of heart failure in young adults. Patients affected with myocarditis can develop dilated cardiomyopathy (DCM), a common reason for heart transplantation, which to date is the only viable option for combatting DCM. Myocarditis/DCM patients show antibodies to coxsackievirus B (CVB)3 and cardiac antigens, suggesting a role for CVB-mediated autoimmunity in the disease pathogenesis; however, a direct causal link remains to be determined clinically. Experimentally, myocarditis can be induced in susceptible strains of mice using the human isolates of CVB3, and the disease pathogenesis of postinfectious myocarditis resembles that of human disease, making the observations made in animals relevant to humans. In this review, we discuss the complex nature of CVB3-induced myocarditis as it relates to the damage caused by both the virus and the host's response to infection. Based on recent data we obtained in the mouse model of CVB3 infection, we provide evidence to suggest that CVB3 infection accompanies the generation of cardiac myosin-specific CD4 T cells that can transfer the disease to naïve recipients. The therapeutic implications of these observations are also discussed.     

Identification of a second mimicry epitope from Acanthamoeba castellanii that induces CNS autoimmunity by generating cross-reactive T cells for MBP 89-101 in SJL mice

We had previously reported that Acanthamoeba castellanii (ACA) contains a mimicry epitope for proteolipid protein 139-151 capable of inducing central nervous system (CNS) autoimmunity in SJL/J mice. We now present evidence that ACA also contains a mimicry epitope for myelin basic protein (MBP) 89-101, a derivative from amoebic nicotinamide adenine dinucleotide dehydrogenase subunit 2 (NAD). The epitope, NAD 108-120, contains a discontinuous stretch of six amino acids in the core region (VVFFKNIILIGFL) sharing 46% identity with MBP 89-101 (VHFFKNIVTPRTP; identical residues are underlined). SJL mice immunized with NAD 108-120 develop encephalomyelitis similar to the disease induced by the cognate peptide. We demonstrate that NAD 108-120 induces T cells that cross-react with MBP 89-101; the antigen-sensitized T cells, which produce predominantly T helper (T(h)) 1 and T(h)17 cytokines, transfer disease in naive SJL recipients reminiscent of the disease induced with MBP 89-101. This is the first report to demonstrate that a solitary microbe can induce CNS autoimmunity by generating cross-reactive T cells for multiple myelin antigens.     

Inhibiting protein phosphatase 2A increases the antitumor effect of protein arginine methyltransferase 5 inhibition in models of glioblastoma

BackgroundDespite multi-model therapy of maximal surgical resection, radiation, chemotherapy, and tumor-treating fields, the median survival of glioblastoma (GBM) patients is less than 15 months. Protein arginine methyltransferase 5 (PRMT5) catalyzes the symmetric dimethylation of arginine residues and is overexpressed in GBM. Inhibition of PRMT5 causes senescence in stem-like GBM tumor cells. LB100, a first-in-class small molecular inhibitor of protein phosphatase 2A (PP2A), can sensitize therapy-resistant tumor cells. Here, we tested the anti-GBM effect of concurrent PRMT5 and PP2A inhibition.MethodsPatient-derived primary GBM neurospheres (GBMNS), transfected with PRMT5 target-specific siRNA, were treated with LB100 and subjected to in vitro assays including PP2A activity and western blot. The intracranial mouse xenograft model was used to test the in vivo antitumor efficacy of combination treatment.ResultsWe found that PRMT5 depletion increased PP2A activity in GBMNS. LB100 treatment significantly reduced the viability of PRMT5-depleted GBMNS compared to PRMT5-intact GBMNS. LB100 enhanced G1 cell cycle arrest induced by PRMT5 depletion. Combination therapy also increased the expression of phospho-MLKL. Necrostatin-1 rescued PRMT5-depleted cells from the cytotoxic effects of LB100, indicating that necroptosis caused the enhanced cytotoxicity of combination therapy. In the in vivo mouse tumor xenograft model, LB100 treatment combined with transient depletion of PRMT5 significantly decreased tumor size and prolonged survival, while LB100 treatment alone had no survival benefit.ConclusionOverall, combined PRMT5 and PP2A inhibition had significantly greater antitumor effects than PRMT5 inhibition alone.     

Recombinant OMP28 antigen-based indirect ELISA for serodiagnosis of bovine brucellosis

Brucellosis is a zoonosis of both public health and economic importance in many developing countries including India. Early detection and segregation of the infected animals are important in order to control the disease. Serodiagnostic tests for brucellosis is mainly based on detection of antibodies developed against lipopolysaccharide (LPS) component of cell. In this study we evaluated a protein antigen, 28 kDa outer membrane protein (OMP28), of Brucella melitensis as an alternative to LPS. Recombinant OMP28 was produced in Escherichia coli system. The efficacy of purified OMP28 was studied in an indirect enzyme-linked immunosorbent assay (ELISA) for diagnosis of brucellosis in field sera collected from different regions of country. Using known negative and known positive serum samples it was found that OMP28 is immunoreactive to Brucella infected cattle, sheep, goat and dog sera. Three hundred and eighty two cattle sera were screened by OMP28 antigen-based ELISA and the results were compared to rose Bengal plate agglutination Test (RBPT). Recombinant OMP28 antigen-based ELISA has shown sensitivity of 88.7%, specificity of 93.8% and accuracy of 92.9%. It was concluded that recombinant B. melitensis OMP28 could be used as a protein antigen for diagnosis of brucellosis in domestic animals.     

Coxsackievirus B3 infection leads to the generation of cardiac myosin heavy chain-α-reactive CD4 T cells in A/J mice

Enteroviruses like coxsackievirus B3 (CVB3) are common suspects in myocarditis/dilated cardiomyopathy patients. Autoimmunity has been proposed as an underlying mechanism, but direct evidence of its role is lacking. To delineate autoimmune response in CVB3 myocarditis, we used IA(k) dextramers for cardiac myosin heavy chain (Myhc)-α 334-352. We have demonstrated that myocarditis-susceptible A/J mice infected with CVB3 generate Myhc-α-reactive CD4 T cells and such a repertoire was absent in na?ve mice as measured by proliferative response to Myhc-α 334-352 and IA(k) dextramer staining. We also detected Myhc-α 334-352 dextramer(+) cells in the hearts of CVB3-infected mice. The autoreactive T cell repertoire derived from infected mice contained a high frequency of interleukin-17-producing cells capable of inducing myocarditis in na?ve recipients. The data suggest that CVB3, a bona fide pathogen of cardiovascular system that primarily infects the heart can lead to the secondary generation of autoreactive T cells and contribute to cardiac pathology.