GGCX mutations show different responses to vitamin K thereby determining the severity of the hemorrhagic phenotype in VKCFD1 patients.

BACKGROUND: Vitamin K dependent coagulation factor deficiency type 1 (VKCFD1) is a rare hereditary bleeding disorder caused by mutations in γ-glutamyl carboxylase (GGCX). VKCFD1 patients are treated life-long with high doses of vitamin K in order to correct the bleeding phenotype. However, normalization of clotting factor activities cannot be achieved for all VKCFD1 patients. OBJECTIVE: The current study aims to investigate the responsiveness to vitamin K for all reported GGCX mutations with respect to clotting factors in order to optimize treatment. METHODS: This study developed an assay using genetically engineered GGCX-/- cells, in which GGCX mutations were analyzed with respect to their ability to γ-carboxylate vitamin K dependent pro-coagulatory and anti-coagulatory clotting factors by ELISA. Additionally, factor VII activity was measured in order to proof protein functionality. For specific GGCX mutations immunofluorescent staining was performed to assess the intracellular localization of clotting factors with respect to GGCX wild-type and mutations. RESULTS: All GGCX mutations were categorized into responder and low responder mutations, thereby determining the efficiency of vitamin K supplementation. Most VKCFD1 patients have at least one vitamin K responsive GGCX allele that is able to γ-carboxylate clotting factors. In few patients, the hemorrhagic phenotype cannot be reversed by vitamin K administration because GGCX mutations on both alleles affect either structural or catalytically important sites thereby resulting in residual ability to γ-carboxylate clotting factors. CONCLUSION: With these new functional data we can predict the hemorrhagic outcome of each VKCFD1 genotype, thus recommending treatments with either vitamin K or prothrombin complex concentrate.

Irgm2 and Gate-16 put a break on caspase-11 activation.

In infections caused by gram-negative bacteria, the bacterial cell wall component lipopolysaccharide (LPS) acts as a potent pathogen-associated molecular pattern (PAMP) that triggers the innate immune system. This is accomplished by two pattern recognition receptor systems. Toll-like receptor 4 (TLR4) senses extracellular LPS and induces a broad pro-inflammatory transcriptional program and also antiviral interferons. A complementary system detects intracellular LPS. As such, upon its release into the cytoplasm, LPS can directly engage the protease caspase-4 (caspase-11 in the murine system) and thereby trigger a pro-inflammatory cell death program known as pyroptosis (Rathinam et al, 2019). This is mediated by active caspase-4 cleaving its substrate gasdermin D (GSDMD). The thereby released N-terminal fragment of GSDMD inserts into the cell membrane and forms a cytotoxic pore. As a consequence, the cell ruptures and releases its pro-inflammatory content. In addition, the GSDMD pore results in potassium efflux that can activate the NLRP3 inflammasome. NLRP3 in turn activates caspase-1, which matures pro-IL-1ß and pro-IL-18, further perpetuating the inflammatory nature of this cell death. Given its unconventional mode of NLRP3 activation, this pathway has been coined the non-canonical inflammasome.

Activation of the NLRP3 Inflammasome by Hyaboron, a New Asymmetric Boron-Containing Macrodiolide from the Myxobacterium Hyalangium minutum.

A Natural Compound Library containing myxobacterial secondary metabolites was screened in murine macrophages for novel activators of IL-1ß maturation and secretion. The most potent of three hits in total was a so far undescribed metabolite, which was identified from the myxobacterium Hyalangium minutum strain Hym3. While the planar structure of 1 was elucidated by high resolution mass spectrometry and NMR data yielding an asymmetric boron containing a macrodiolide core structure, its relative stereochemistry of all 20 stereocenters of the 42-membered ring was assigned by rotating frame Overhause effect spectroscopy correlations, 1H,1H, and 1H,13C coupling constants, and by comparison of 13C chemical shifts to those of the structurally related metabolites tartrolon B-D. The absolute stereochemistry was subsequently assigned by Mosher's and Marfey's methods. Further functional studies revealed that hyaboron and other boronated natural compounds resulted in NLRP3 inflammasome dependent IL-1ß maturation, which is most likely due to their ability to act as potassium ionophores. Moreover, besides its inflammasome-stimulatory activity in human and mouse cells, hyaboron (1) showed additional diverse biological activities, including antibacterial and antiparasitic effects.