Inefficient and abortive classical complement pathway activation by the calcium inositol hexakisphosphate component of the Echinococcus granulosus laminated layer

Persistent extracellular tissue-dwelling pathogens face the challenge of antibody-dependent activation of the classical complement pathway (CCP). A prime example of this situation is the larva of the cestode Echinococcus granulosus sensu lato, causing cystic echinococcosis. This tissue-dwelling, bladder-like larva is bounded by a cellular layer protected by the outermost acellular “laminated layer” (LL), to which host antibodies bind. The LL is made up of a mucin meshwork and interspersed nano-deposits of calcium inositol hexakisphosphate (calcium InsP₆). We previously reported that calcium InsP₆ bound C1q, apparently initiating CCP activation. The present work dissects CCP activation on the LL. Most of the C1 binding activity in the LL corresponded to calcium InsP₆, and this binding was enhanced by partial proteolysis of the mucin meshwork. The remaining C1 binding activity was attributable to host antibodies, which included CCP-activating IgG isotypes. Calcium InsP₆ made only a weak contribution to early CCP activation on the LL, suggesting inefficient C1 complex activation as reported for other polyanions. CCP activation on calcium InsP₆ gave rise to a dominant population of C3b deposited onto calcium InsP₆ itself that appeared to be quickly inactivated. Apparently as a result of inefficient initiation plus C3b inactivation, calcium InsP₆ made no net contribution to C5 activation. We propose that the LL protects the underlying parasite cells from CCP activation through the combined effects of inefficient permeation of C1 through the mucins and C1 retention on calcium InsP₆. This mechanism does not result in C5 activation, which is known to drive parasite-damaging inflammation.     

Monoclonal antibody against human Tim‐3 enhances antiviral immune response

T cell immunoglobulin and mucin domain protein 3 (Tim‐3) is an immune checkpoint inhibitor in T cells and innate immune cells. The deregulated upregulation of Tim‐3 is related to immune exhaustion in tumour and viral infection. To overcome Tim‐3‐mediated immune tolerance, we developed a novel monoclonal antibody against human Tim‐3 (L3G) and investigated its roles in inhibiting Tim‐3 signalling and overcoming immune tolerance in T cells and monocytes/macrophages. The administration of L3G to cultured peripheral blood mononuclear cells (PBMCs) significantly increased the production of IFN‐γ and IL‐2 and the expression of type I interferon. The administration of L3G also increased the production of IFN‐γ, IL‐8 and type I interferon in U937 cells and primary monocytes. We investigated the mechanisms by which L3G enhances pro‐inflammatory cytokine expression, and our data show that L3G enhances STAT1 phosphorylation in both monocytes/macrophages and T cells. Finally, in an H1N1 infection model of PBMCs and U937 cells, L3G decreased the viral load and enhanced the expression of interferon. Thus, we developed a functional antibody with therapeutic potential against Tim‐3‐mediated infection tolerance.     

Construction and characterization of an OmpH-deficient mutant of Pasteurella multocida strain X-73

A capsule-defective mutant strain PBA129 of Pasteurella multocida was constructed by electroporation of phagemid containing the coding region of the antisense RNA of the ompH gene into the wild type strain X-73 (serovar A:1) of P. multocida. The pathogenicity and protective potency of the mutant against homologous and heterologous challenge in mice and chickens were characterized. Greyish colonies of the mutant, indicating lower capsule thickness, on selective dextrose starch agar were observed under an obliquely transmitted light stereomicroscope and compared to iridescent colonies of the wild type strain X-73. Strain PBA129 had lower capsule thickness than the wild type strain as observed with an electron microscope. Strain PBA129 was apparently attenuated, as mice and chickens inoculated with the bacteria at 10⁸ CFU survived. Protection was observed in both mice and chickens inoculated with strain PBA129 upon challenge exposure to avian P. multocida strains X-73 and P-1059 (serovar A:3), respectively. In conclusion, the mutant strain PBA129 of P. multocida strain X-73 was completely attenuated, and it was possible to induce sufficient protection against avian P. multocida strains.     

Proteases and their inhibitors as prognostic factors for high-grade serous ovarian cancer

Ovarian carcinoma is one of the most lethal malignancies, but only very few prognostic biomarkers are known. The degradome, comprising proteases, protease non-proteolytic homologues and inhibitors, have been involved in the prognosis of many cancer types, including ovarian carcinoma. The prognostic significance of the whole degradome family has not been specifically studied in high-grade serous ovarian cancer. A targeted DNA microarray known as the CLIP-CHIP microarray was used to identify potential prognostic factors in ten high-grade serous ovarian cancer women who had early recurrence (<1.6 years) or late/no recurrence after first line surgery and chemotherapy. In women with early recurrence, we identified seven upregulated genes (TMPRSS4, MASP1/3, SPC18, PSMB1, IGFBP2, CFI – encoding Complement Factor I – and MMP9) and one down-regulated gene (ADAM-10). Using immunohistochemistry, we evaluated the prognostic effect of these 8 candidate genes in an independent cohort of 112 high-grade serous ovarian cancer women. Outcomes were progression, defined according to CA-125 criteria, and death. Multivariate Cox proportional hazard regression models were done to estimate the associations between each protein and each outcome. High ADAM-10 expression (intensity of 2–3) was associated with a lower risk of progression (adjusted hazard ratio (HR): 0.51; 95% confidence interval (CI): 0.29-0.87). High complement factor I expression (intensity 2–3) was associated with a higher risk of progression (adjusted HR: 2.30, 95% CI: 1.17–4.53) and death (adjusted HR: 3.42; 95% CI: 1.72–6.79). Overall, we identified the prognostic value of two proteases, ADAM-10 and complement factor I, for high-grade serous ovarian cancer which could have clinical significance.     

A fluorescence method for homogeneous detection of influenza A DNA sequence based on guanine-quadruplex-N-methylmesoporphyrin IX complex and assistance-DNA inhibition

In his study, we report a fluorescence method for homogeneous detection of influenza A (H1N1) DNA sequence based on G-quadruplex-NMM complex and assistance-DNA (A-DNA) inhibition. The quadruplex-based functional DNA (QBF-DNA), composed of a complementary probe to the target H1N1 DNA sequence and G-rich fragment, was designed as the signal DNA. The A-DNA consisted of two parts, one part was complementary to target H1N1 DNA and the other part was complementary to the signal DNA. In the absence of target H1N1 DNA, the G-rich fragment of QBF-DNA can form G-quadruplex-NMM complex, which outputted a fluorescent signal. With the presence of target H1N1 DNA, QBF-DNA, and A-DNA can simultaneously hybridize with target H1N1 DNA to form double-helix structure. In this case, the A-DNA partially hybridized with the QBF-DNA, which inhibited the formation of G-quadruplex-NMM complex, leading to the decrease of fluorescent signal. Under the optimum conditions, the fluorescence intensity was inversely proportional to the concentration of target H1N1 DNA over the range from 25 to 700 pmol/L with a detection limit of 8 pmol/L. In addition, the method is target specific and practicability, and would become a new diagnostic assay for H1N1 DNA sequence and other infectious diseases.     

Macular buckle without vitrectomy for myopic macular schisis: a Canadian case series

Objective

To determine the effectiveness of a macular buckle procedure without vitrectomy for the treatment of symptomatic myopic macular schisis.

Should we look for Hirschsprung disease in all children with meconium plug syndrome?

BackgroundMeconium plug syndrome (MPS) is associated with Hirschsprung disease (HD) in 13–38% of cases. This study sought to assess institutional variation in utilization of rectal biopsy in children with MPS and the likelihood of diagnosing HD in this population.MethodsPatients with MPS on contrast enema in the first 30 days of life from the Pediatric Health Information System database in 2016–2017 were included. Institutional rates of rectal biopsies performed during the initial admission were calculated and then used to predict institutional rates of early HD diagnoses using Poisson regression.ResultsOf 373 newborns with MPS, 106 (28.4%) underwent early rectal biopsy, of whom 43 (40.5%) had HD. Fifty-seven (15.3%) were ultimately diagnosed with HD. Eight (14%) of these patients had a delayed diagnosis. HD rates between institutions did not differ significantly (range 0–50%, p = 0.52), but usage of early rectal biopsy did (range 0–80%, p = 0.03). Each additional early biopsy increased the early HD diagnosis rate by 35% (β = 0.30, 95% CI 0.15–0.45, p < 0.0001).ConclusionThe incidence of HD is increased in children with MPS. There is significant hospital variability in the utilization of early rectal biopsy, and opportunity exists to standardize practice.Type of Study: Study of Diagnostic testLevel of Evidence: Level III