Vibrio MARTX toxin processing and degradation of cellular Rab GTPases by the cytotoxic effector Makes Caterpillars Floppy.

Vibrio vulnificus causes life-threatening wound and gastrointestinal infections, mediated primarily by the production of a Multifunctional-Autoprocessing Repeats-In-Toxin (MARTX) toxin. The most commonly present MARTX effector domain, the Makes Caterpillars Floppy-like (MCF) toxin, is a cysteine protease stimulated by host adenosine diphosphate (ADP) ribosylation factors (ARFs) to autoprocess. Here, we show processed MCF then binds and cleaves host Ras-related proteins in brain (Rab) guanosine triphosphatases within their C-terminal tails resulting in Rab degradation. We demonstrate MCF binds Rabs at the same interface occupied by ARFs. Moreover, we show MCF preferentially binds to ARF1 prior to autoprocessing and is active to cleave Rabs only subsequent to autoprocessing. We then use structure prediction algorithms to demonstrate that structural composition, rather than sequence, determines Rab target specificity. We further determine a crystal structure of aMCF as a swapped dimer, revealing an alternative conformation we suggest represents the open, activated state of MCF with reorganized active site residues. The cleavage of Rabs results in Rab1B dispersal within cells and loss of Rab1B density in the intestinal tissue of infected mice. Collectively, our work describes an extracellular bacterial mechanism whereby MCF is activated by ARFs and subsequently induces the degradation of another small host guanosine triphosphatase (GTPase), Rabs, to drive organelle damage, cell death, and promote pathogenesis of these rapidly fatal infections.

A Genomic Island of Vibrio cholerae Encodes a Three-Component Cytotoxin with Monomer and Protomer Forms Structurally Similar to Alpha-Pore-Forming Toxins.

Alpha-pore-forming toxins (α-PFTs) are secreted by many species of bacteria, including Escherichia coli, Aeromonas hydrophila, and Bacillus thuringiensis, as part of their arsenal of virulence factors, and are often cytotoxic. In particular, for α-PFTs, the membrane-spanning channel they form is composed of hydrophobic α-helices. These toxins oligomerize at the surface of target cells and transition from a soluble to a protomer state in which they expose their hydrophobic regions and insert into the membrane to form a pore. The pores may be composed of homooligomers of one component or heterooligomers with two or three components, resulting in bi- or tripartite toxins. The multicomponent α-PFTs are often expressed from a single operon. Recently, motility-associated killing factor A (MakA), an α-PFT, was discovered in Vibrio cholerae. We report that makA is found on the V. cholerae GI-10 genomic island within an operon containing genes for two other potential α-PFTs, MakB and MakE. We determined the X-ray crystal structures for MakA, MakB, and MakE and demonstrated that all three are structurally related to the α-PFT family in the soluble state, and we modeled their protomer state based on the α-PFT AhlB from A. hydrophila. We found that MakA alone is cytotoxic at micromolar concentrations. However, combining MakA with MakB and MakE is cytotoxic at nanomolar concentrations, with specificity for J774 macrophage cells. Our data suggest that MakA, -B, and -E are α-PFTs that potentially act as a tripartite pore-forming toxin with specificity for phagocytic cells. IMPORTANCE The bacterium Vibrio cholerae causes gastrointestinal, wound, and skin infections. The motility-associated killing factor A (MakA) was recently shown to be cytotoxic against colon, prostate, and other cancer cells. However, at the outset of this study, the capacity of MakA to damage cells in combination with other Mak proteins encoded in the same operon had not been elucidated. We determined the structures of three Mak proteins and established that they are structurally related to the α-PFTs. Compared to MakA alone, the combination of all three toxins was more potent specifically in mouse macrophages. This study highlights the idea that the Mak toxins are selectively cytotoxic and thus may function as a tripartite toxin with cell type specificity.

Deep Learning in Medical Hyperspectral Images: A Review.

With the continuous progress of development, deep learning has made good progress in the analysis and recognition of images, which has also triggered some researchers to explore the area of combining deep learning with hyperspectral medical images and achieve some progress. This paper introduces the principles and techniques of hyperspectral imaging systems, summarizes the common medical hyperspectral imaging systems, and summarizes the progress of some emerging spectral imaging systems through analyzing the literature. In particular, this article introduces the more frequently used medical hyperspectral images and the pre-processing techniques of the spectra, and in other sections, it discusses the main developments of medical hyperspectral combined with deep learning for disease diagnosis. On the basis of the previous review, tne limited factors in the study on the application of deep learning to hyperspectral medical images are outlined, promising research directions are summarized, and the future research prospects are provided for subsequent scholars.

A state-of-the-science review and guide for measuring environmental exposure biomarkers in dried blood spots.

BACKGROUND: Dried blood spot (DBS) sampling is a simple, cost-effective, and minimally invasive alternative to venipuncture for measuring exposure biomarkers in public health and epidemiological research. DBS sampling provides advantages in field-based studies conducted in low-resource settings and in studies involving infants and children. In addition, DBS samples are routinely collected from newborns after birth (i.e., newborn dried blood spots, NDBS), with many states in the United States permitting access to archived NDBS samples for research purposes. OBJECTIVES: We review the state of the science for analyzing exposure biomarkers in DBS samples, both archived and newly collected, and provide guidance on sample collection, storage, and blood volume requirements associated with individual DBS assays. We discuss recent progress regarding analytical methods, analytical sensitivity, and specificity, sample volume requirements, contamination considerations, estimating extracted blood volumes, assessing stability and analyte recovery, and hematocrit effects. METHODS: A systematic search of PubMed (MEDLINE), Embase (Elsevier), and CINAHL (EBSCO) was conducted in March 2022. DBS method development and application studies were divided into three main chemical classes: environmental tobacco smoke, trace elements (including lead, mercury, cadmium, and arsenic), and industrial chemicals (including endocrine-disrupting chemicals and persistent organic pollutants). DBS method development and validation studies were scored on key quality-control and performance parameters by two members of the review team. RESULTS: Our search identified 47 published reports related to measuring environmental exposure biomarkers in human DBS samples. A total of 28 reports (37 total studies) were on methods development and validation and 19 reports were primarily the application of previously developed DBS assays. High-performing DBS methods have been developed, validated, and applied for detecting environmental exposures to tobacco smoke, trace elements, and several important endocrine-disrupting chemicals and persistent organic pollutants. Additional work is needed for measuring cadmium, arsenic, inorganic mercury, and bisphenol A in DBS and NDBS samples. SIGNIFICANCE: We present an inventory and critical review of available assays for measuring environmental exposure biomarkers in DBS and NDBS samples to help facilitate this sampling medium as an emerging tool for public health (e.g., screening programs, temporal biomonitoring) and environmental epidemiology (e.g., field-based studies).

An Ovarian Steroid Metabolomic Pathway Analysis in Basal and Polycystic Ovary Syndrome (PCOS)-like Gonadotropin Conditions Reveals a Hyperandrogenic Phenotype Measured by Mass Spectrometry.

Prior work has demonstrated that murine ovarian explants and isolated ovarian follicles can recapitulate human-like 28-day cycles in vitro with normal patterns of estradiol and progesterone secretion in response to gonadotropin stimulation. The objective of this study was to manipulate the gonadotropin stimulation protocol to mimic polycystic ovary syndrome (PCOS) and assess the resulting changes in ovarian steroidogenesis. A secondary aim of the study was to develop a high-throughput, sensitive, and specific liquid chromatography with tandem mass spectrometry (LC-MS/MS) assay to measure seven steroid hormones (estrone, estradiol, progesterone, testosterone, androstenedione, dehydroepiandrosterone, and dihydrotestosterone) in conditioned culture media. Ovaries were harvested from 12-day-old CD-1 mice and cultured for 28 days, with ovulation induction on culture day 14. Media were supplemented human chorionic gonadotropin (hCG, a luteinizing hormone analog) and follicle stimulating hormone (FSH) at ratios of 1:0 (standard media), 1:1 (physiologic ratio), and 3:1 (PCOS-like ratio). Ovaries cultured in PCOS-like media displayed hyperandrogenism and impaired ovulation, two key features of a PCOS-like phenotype. Taken together, this first-of-its-kind presentation of hormone levels from single tissues creates a map of the enzymatic steps most acutely affected by gonadotropin dysregulation and may provide opportunities for assessing other potential insults in PCOS pathogenesis.

Human Serum Albumin Cys34 Adducts in Newborn Dried Blood Spots: Associations With Air Pollution Exposure During Pregnancy.

Background: Increasing evidence suggests that exposure to air pollution during pregnancy is associated with adverse pregnancy outcomes. However, biomarkers associated with air pollution exposure are widely lacking and often transient. In addition, ascertaining biospecimens during pregnacy to assess the prenatal environment remains largely infeasible. Objectives: To address these challenges, we investigated relationships between air pollution exposure during pregnancy and human serum albumin Cys34 (HSA-Cys34) adducts in newborn dried blood spots (DBS) samples, which captures an integration of perinatal exposures to small reactive molecules in circulating blood. Methods: Newborn DBS were obtained from a state archive for a cohort of 120 children born at one Kaiser Permanente Southern California (KPSC) hospitals in 2007. These children were selected to maximize the range of residential air pollution exposure during the entire pregnancy to PM2.5, PM10, NO2, O3, based on monthly estimates interpolated from regulatory monitoring sites. HSA-Cys34 adducts were selected based on previously reported relationships with air pollution exposure and oxidative stress. Results: Six adducts measured in newborn DBS samples were associated with air pollution exposures during pregnancy; these included direct oxidation products, adducts formed with small thiol compounds, and adducts formed with reactive aldehydes. Two general trends were identified: Exposure to air pollution late in pregnancy (i.e., in the last 30 days) was associated with increased oxidative stress, and exposure to air pollution earlier in pregnancy (i.e., not in the last 30 days) was associated with decreased oxidative stress around the time of birth. Discussion: Air pollution exposure occurring during pregnancy can alter biology and leave measurable impacts on the developing infant captured in the newborn DBS adductome, which represents a promising tool for investigating adverse birth outcomes in population-based studies.