Tissue Sections Research Articles

First Report of Rhizoctonia solani AG 4 HG-III Causing Stem Lesion of Watermelon in Idaho.

In July 2022, stem lesions, approximately 4 to 5 cm in length as well as leaf wilt and dark brown necrosis on stems and roots were observed in two fields in Southwest Idaho on 20 to 30% of watermelons (Citrullus lanatus). To determine the causal agent, isolations were attempted from symptomatic tissue. The surface of the affected material was disinfected with 0.6% sodium hypochlorite for 1 min and rinsed three times with sterile water. Approximately 2 mm3 sections of tissue were plated on water agar amended with 0.02% penicillin and 0.08% streptomycin and incubated at room temperature for 7 days. Fungal colonies were tentatively identified as Rhizoctonia from right-angle branching and septate hyphal structures, slight constriction and septum near the branch base, and the production of 1 to 2 mm white to light brown irregularly shaped sclerotia. Single hyphal tips were transferred to potato dextrose agar (PDA) and grown at room temperature. Approximately ten isolates from each field with a consistent macromorphology were observed. These isolates had light brown mycelia, produced sclerotia at ambient temperature with no exposure to continuous light, and a representative isolate, designated D22-110 was selected for sequencing and pathogenicity testing. For isolate D22-110, mycelia were removed with a scalpel after 7 days of growth, for DNA extraction and sequencing of the rDNA internal transcribed spacer (ITS) region as previously described (White et al., 1990). A 726 bp product was generated and the sequence was submitted to GenBank (Accession No. OQ794049). NCBI-BLAST indicated this sequence was 99% identical (631 of 634 bp and 632 of 634 bp identical) with known reference isolates previously identified as R. solani AG 4 HG-III (Accession No. AF354075 and AF354076, respectively) from a phylogenetic study (Gonzalez et al., 2001). Pathogenicity testing was performed twice on two-week-old seedlings of watermelon cultivars Endless Summer and Wingman in greenhouse conditions (29oC, 12 h daylight). Two disks (3 mm diam) from 7-day-old plates of PDA were placed around each seedling at the root and stem convergence point. Ten seedlings were mock-inoculated with sterile PDA plugs as a control. Approximately 35% damping-off incidence was observed on inoculated seedlings six days post-inoculation, while control seedlings remained healthy. At 20 days post-inoculation, 20 (first trial) and 34 seedlings (second trial) were assessed for visible stem and root lesions. Incidence of stem lesions occurred on 90% of seedlings, with 80% of seedlings possessing lesions greater than 10 mm in diameter. Seedlings without R. solani inoculation were free of stem and root lesions. R. solani was re-isolated from symptomatic tissue, with 40% frequency of isolation, identified by right-angle branching of the hyphae thus confirming Koch's postulates. R. solani AG 4 has been reported in watermelon in the US since 1994 (Hall and Summer, 1994) but the AG 4 subgroup was not reported. AG 4 HG-III was reported in melon seedlings causing damping-off in Kyrgyzstan (Erper et al. 2016). In other hosts, AG 4 HG-III was found in potatoes in South Africa (Muzhinji et al., 2014), buckwheat and foxtail millet in China (Zhou et al., 2015; Hao et al., 2023), broccoli and spinach (Kuramae et al., 2003) and turnip green (Sekiguchi et al., 2015). To the best of our knowledge, this is the first report of R. solani AG 4 HG-III causing disease in watermelon in Idaho. Given the the rate of disease incidence observed in the field, growers should consider avoiding planting alternative host crops to minimize inoculum buildup.

Effects of 17α-methyltestosterone and letrozole on growth and gonadal development in largemouth bass (Micropterus salmodies)

In order to optimize the parameters for reversing masculinization and establish the techniques for sex induction of pseudo-males and creation of all-female fry in largemouth bass (Micropterus salmodies, LMB), 15-day-old LMB (1.00 ± 0.10 cm in length, 0.10 ± 0.01 g in weight) were fed a diet supplemented with either 17α-methyltestosterone (MT) or letrozole (LE) and their combination. The experimental groups were M20 (20 mg/kg MT), L20 (20 mg/kg LE) and M10L10 (10 mg/kg MT and 10 mg/kg LE). The control group, named C, was not feed MT or LE. After 60 days, exogenous hormone in the diets was stopped and the effects of MT and LE on growth, male ratio, and gonadal development in LMB were evaluated. At 12-month-old, blood and gonadal tissue samples were collected to measure sex steroid hormones levels, analyze expression levels of dmrt1 and cyp19a1a genes, as well as examine the gonads morphology. The results showed no significant differences in growth between the experimental groups and the control group after a 60-day feeding period with the formulated diet (p > 0.05). The sex reversal ratio of M20, L20, M10L10 were 95.00%, 80.00%, 76.47%, respectively. The gonadal tissue sections showed that the gonadal structure of masculinized fish morphologic resembled that of control male fish. At 12-month-old, the sex reversal ratio in M20, L20, M10L10 and C groups were 100%, 86.67%, 73.33% and 50.00%, respectively. The testicular of pseudo-male fish in the M20 group exhibited well-developed morphology similarities to that of the control group males. However, the testes of pseudo-male fish in the L20 and M10L10 groups were smaller size Estradiol (E2) levels in the experimental groups was significantly lower than those in the control group females (p < 0.05), while testosterone (T) levels were significantly higher than that of the control group (p < 0.05). Compared to the female fish in the control group, pseudo-male fish from all experimental groups showed significantly upregulated expression of dmrt1 (p < 0.05), and significantly downregulated expression of cyp19a1a (p < 0.05). Pseudo-males selected from group M20 exhibited a significantly higher proportion of female offspring (92.00%) compared to the control group (46.50%). In summary, 20 mg/kg MT was the optimal inducing concentration.

Use of Melinex film for flat embedding tissue sections in LR White.

Tissue slices can undergo distortions during processing into resin for light and electron microscopy as a result of differential shrinkage of the various tissue components, and this may necessitate removal of a considerable amount of material from the final resin-embedded tissue block to ensure production of complete sections of the sample. To mitigate this problem, a number of techniques have been devised that ensure the sample is held flat during the final curing/polymerisation of the resin. For embedding in acrylic resins, oxygen must be excluded as it inhibits polymerisation, and methods devised for epoxy resin embedding are generally unsuitable. The method describes the preparation and use of air-tight flat-embedding chambers prepared from Melinex film and provides an inexpensive, technically simpler, and versatile alternative to chambers formed from either Thermanox coverslips or Aclar films that have previously been advocated for such purposes. Lay description: Tissue slices can undergo distortions during processing into resin for light and electron microscopy as a result of differential shrinkage of the various tissue components. Such distortions may necessitate removal of a considerable amount of material to ensure production of complete sections of the sample. For embedding in acrylic resins, oxygen must be excluded as it inhibits polymerisation, and methods devised for epoxy resin flat-embedding are generally unsuitable. Air-tight flat-embedding chambers prepared from either Thermanox coverslips, or a combination of PTFE-coated glass slides, polycarbonate film gaskets, and Aclar film have been advocated for such purposes. Thermanox coverslips are expensive and limited in size to 22mm × 60mm, and the alternative method is technically complicated. Melinex film is commercially available as 210mm × 297mm sheets and is approximately 1/20th the price of Thermanox and less than half the price of Aclar film. The method describes the preparation and use of embedding chambers made from Melinex film, glass slides and double-sided adhesive tape as a technically simpler, inexpensive and versatile alternative to both Thermanox coverslips and the Aclar film method.

Abstract PR-13: Disruption of sulfatide metabolism by targeting UGT8 is an actionable metabolic vulnerability for pancreatic cancer early interception

Abstract The existence of asymptomatic precursor lesions that are known to predate invasive pancreatic ductal adenocarcinoma (PDAC) by years provides a compelling rationale for cancer interception efforts. One such precursor is intraductal papillary mucinous neoplasm (IPMN). Using Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry (MALDI-MS) imaging of metabolites in tissue sections from resected patient IPMN cases (N=23) and from a KrasG12D/GnasR201C mouse model of IPMN, we have made a discovery of long chain hydroxylated sulfatide species as being selectively enriched in IPMN. Spatial transcriptomics analysis of serial sections from the same IPMN tissues confirmed cognate transcripts, including ceramide galactosyltransferase (CGT, also known as UGT8) and galactose-3-O-sulfotransferase (CST, also known as Gal3St1) enzymes that, respectively, catalyze the addition of galactose from UDP-galactose to ceramide and the O-sulfation of the galactose residue on galactosylceramide (GalCer), to also be highly enriched in IPMN. Functional studies using CRISPR/cas9 UGT8 knockout Kras;Gnas IPMN cells demonstrated that loss of UGT8, the key enzyme that produces the sulfatide precursor GalCer, results in attenuation of sulfatide biosynthesis and impaired mitochondrial function, as evidenced by reduced mitochondrial respiration and ATP production. These changes were met with concomitant decreases in proliferation and invasiveness, and increased susceptibility to intrinsic apoptosis. Targeting of sulfatide metabolism using a selective small molecule inhibitor of UGT8 suppressed sulfatide biosynthesis, induced ceramide-mediated compensatory mitophagy, and subsequent apoptosis in Kras;Gnas IPMN cells. In vivo, UGT8 inhibition suppressed tumor growth in an IPMN allograft model. In conclusion, our work identifies enhanced sulfatide metabolism as an early metabolic alteration in cystic pre-cancerous lesions of the pancreas that persists through invasive neoplasia and represents an actionable vulnerability for PDAC early interception. Citation Format: Riccardo Ballarò, Yihui Chen, Fredrik I Thege, Rongzhang Dou, Jimin Min, Michele Yip-Schneider, Jianjun Zhang, Ranran Wu, Ehsan Irajizad, Yuki Makino, Kimal I Rajapakshe, Mark W Hurd, Ricardo A León-Letelier, Edwin Ostrin, Jody Vykoukal, Jennifer B Dennison, Kim-Anh Do, Samir Hanash, Robert A Wolff, Paola A Guerrero, Michael P Kim, C Max Schmidt, Anirban Maitra, Johannes F Fahrmann. Disruption of sulfatide metabolism by targeting UGT8 is an actionable metabolic vulnerability for pancreatic cancer early interception [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr PR-13.

Abstract A046: Spatial transcriptomics reveals heterogeneity of epithelial and stromal compartments in healthy and tumor pancreas

Abstract Pancreatic Ductal Adenocarcinoma (PDAC) is one of the most lethal cancers, with a 5-year survival rate of only 13%. PDAC develops from precursor lesions, most commonly Pancreatic Intraepithelial Neoplasia (PanIN). Studying PanIN in human patients is difficult due to its microscopic nature and the lack of indication for sampling the pancreas in the absence of disease. In collaboration with Gift of Life - Michigan, a local organ and tissue donor organization, we obtained over 80 pancreata for research. Surprisingly, we found PanIN in the vast majority of the organs, regardless of the donor’s age; the prevalence of PanIN compared to the relatively low incidence of PDAC indicates that most lesions are destined not to progress. To gather insight into factors that might distinguish lesions designed to progress from lesions that are likely to remain dormant, we sought to characterize both epithelial cells and their supporting microenvironment. Using spatial transcriptomics, we determined that PanIN epithelial cells are transcriptionaally similar to pancreatic cancer; further, limited genetic characterization revealed that they contain similar oncogenic KRAS mutations as pancreatic malignancy. We then sought to determine whether factors in the microenvironment distinguish sporadic PanIN from those associated with overt malignancy. We used 10x Genomic CytAssist Visium technology and profiled an initial batch of 7 tissue sections from donor pancreata and an equal number from tumors. We employed the BayesPrism algorithm to deconvolute the Visium data (which lacked single-cell resolution), using our single-cell RNAseq atlas to define stroma- and epithelial-specific expression profiles. We then segmented the tissue into epithelial and stromal compartments. We further identified various stromal subpopulations, with macrophages and fibroblasts being the most abundant cell types. We defined subpopulations of epithelial cells, fibroblasts, and macrophages through unsupervised clustering. In both donor samples and tumors, we identified heterogeneous populations of stromal cells present in specific geographic patterns, with PanIN-adjacent stroma clearly distinct from stroma surrounding healthy ducts. Cell-cell neighborhood analysis revealed a distinct geographic arrangement of epithelial and stromal compartments, indicating potential communication patterns. Independent unsupervised statistical analysis using Non-negative Matrix Factorization (NMF) using Coordinated Gene Association in Pattern Sets (CoGAPS) R package also revealed similar patterns. Our next steps involve validating gene expression signatures that define each subpopulation and identifying tumor-specific or PanIN-specific genes. Ultimately, we aim to investigate interactions between epithelial cells, fibroblasts, and macrophages to identify potential tumor-promoting or tumor-restraining pathways. Citation Format: Ahmed M. Elhossiny, Jude Okoye, Imade Williams, Yaqing Zhang, Hannah Watkoske, Atul Deshpande, Elana Fertig, Arvind Rao, Eileen S. Carpenter, Timothy L. Frankel, Marina Pasca di Magliano. Spatial transcriptomics reveals heterogeneity of epithelial and stromal compartments in healthy and tumor pancreas [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A046.

Mass Spectrometry Imaging for the Characterization of C═C Localization in Unsaturated Lipid Isomers at the Single-Cell Level.

Unsaturated lipids with carbon-carbon double bonds (C═C) have been implicated in the pathogenesis of various diseases. While mass spectrometry imaging (MSI) has been employed to map the distribution of lipid isomers in tissue sections, the identification of lipid C═C positional isomers at the single-cell level using MSI poses a significant challenge. In this study, we developed a novel approach utilizing ToF-SIMS in conjunction with the Paternò-Büchi (P-B) photochemical reaction to characterize the C═C localization in unsaturated lipid isomers at the single-cell level. The P-B reaction was employed to produce adduct products, which were subsequently subjected to collision-induced dissociation by the primary ion beam of ToF-SIMS to generate characteristic ion pairs indicative of the presence of C═C bonds. Utilizing this approach, lipid isomers in brain and skeletal tissues from mice, as well as different cell lines, were visualized at single-cell resolution. Furthermore, distinct variations in the composition of FA 18:1 isomers across different microregions and cell types were revealed. Our P-B ToF-SIMS approach enables the accurate identification and characterization of complex lipid structures with remarkable spatial resolution and can be helpful in understanding the physiological role of these C═C positional isomers.

Hyperbranched Poly-l-Lysine Modified Titanium Surface With Enhanced Osseointegration, Bacteriostasis, and Anti-Inflammatory Properties for Implant Application: AnExperimental InVivo Study.

This study aimed to explore multiple effects of hyperbranched poly-l-lysine (HBPL) titanium (Ti) surfaces on osseointegration, bacteriostasis, and anti-inflammation across three different animal models. Ti surfaces were covalently modified with HBPL, with uncoated surfaces as controls. Characterization included scanning electron microscopy (SEM) and surface chemistry and elemental analysis (EDX). Ti and Ti-HBPL implants were placed in conventional canine edentulous sites, post-operative infection canine edentulous sites, and diabetic rat tibias. Implants from canine edentulous models were analyzed using micro-CT and histomorphometry to assess osseointegration at 8 weeks. Post-operative infection beagles were used to evaluate antibacterial efficacy through clinical parameters and bacterial cultures at 1 week. In diabetic rats, micro-CT and histomorphometry were performed at 8 weeks. HBPL was uniformly grafted on Ti-HBPL surfaces. Ti-HBPL surfaces showed higher bone volume/total volume (BV/TV, p < 0.001), bone-implant contact (BIC%, p < 0.001), and trabecular number (Tb.N, p < 0.01) in beagles. Besides, it displayed higher BIC% (p < 0.001) and bone area fraction occupancy (BAFO%, p < 0.01) in hard tissue sections. In an infected model, Ti-HBPL surfaces exhibited lower bleeding on probing (BOP, p < 0.001), and plaque index (DI, p < 0.01), with reduced bacterial colony formation (p < 0.001) compared to the control group. In diabetic rats, Ti-HBPL surfaces showed an increase in BV/TV (p < 0.01) and Tb.N (p < 0.001), downregulated TNF-α and IL-1β (p < 0.01), and upregulated IL-10 (p < 0.01) and osteocalcin (OCN) expression (p < 0.01). HBPL-Ti surfaces demonstrated enhanced osseointegration, bacteriostasis, and anti-inflammatory effects invivo.

Effects of anti-rainbow trout germ-cell monoclonal antibody on germ cells and gonadal tissues in rainbow trout (Oncorhynchus mykiss)

The development of sterilization technology is important to improve the production efficacy of aquaculture and prevent genetically modified fish from escaping. Immunosterilization is a novel sterilization method that has already been used in mammals. In this study, anti-rainbow trout germ-cell monoclonal antibodies (anti-GC mAbs) were injected into immature rainbow trout Oncorhynchus mykiss, and their effects were analyzed. First, anti-GC mAbs labeled with Alexa Fluor 488 were injected into the body cavity of larvae 20 days (group 1) and 102 days (group 2) after fertilization. Second, anti-GC mAbs were injected into the body cavity of yearling trout (group 3), and their impacts on gonadal tissue were analyzed in terms of histology and gene expression levels. As a result, anti-GC mAbs were observed to bind to some primordial GCs in group 1, whereas there were no obvious positive cells in male testes of group 2. In contrast, green fluorescent signals were observed in an arrangement parallel to the ovarian lamina in female ovaries of group 2, which was similar to the arrangement of positive signals on ovarian tissue sections immunostained with anti-GC mAbs. GCs in group 1 were observed again after 76 days, and double-positive cells remained. In group 3, there were changes in the expression levels of GC-specific genes, but the changes were not seen as biologically meaningful. Moreover, there was no histological abnormality in gonadal tissues after immunization. In conclusion, these results suggested that anti-GC mAbs could only reach GCs in early developmental stages or in the ovary, and injection of anti-GCs mAbs did not have harmful effects on GCs after binding.

Gram stain of skin tissue sections: The difficulty in establishing diagnostic standards for skin and soft tissue infections.

Gram stain of skin tissue sections: The difficulty in establishing diagnostic standards for skin and soft tissue infections.

A graph self-supervised residual learning framework for domain identification and data integration of spatial transcriptomics

Spatial transcriptomics (ST) technologies allow for comprehensive characterization of gene expression patterns in the context of tissue microenvironment. However, accurately identifying domains with spatial coherence in both gene expression and histology in situ and effectively integrating data from multi-sample remains challenging. Here, we propose ResST, a graph self-supervised residual learning model based on graph neural network and Margin Disparity Discrepancy (MDD) theory. ResST aggregates gene expression, biological effects, spatial location, and morphological information to capture nonlinear relationships between a cell and surrounding cells for spatial domain identification. Also, ResST integrates multiple ST datasets and aligns latent embeddings based on MDD theory for correcting batch effects. Results show that ResST identifies continuous spatial domains at a finer scale in ten ST datasets acquired with different technologies. Moreover, ResST efficiently integrated data from multiple tissue sections vertically or horizontally while correcting batch effects. Overall, ResST demonstrates exceptional performance in analyzing ST datasets.

Tetrandrine ameliorated atherosclerosis in vitamin D3/high cholesterol diet-challenged rats via modulation of miR-34a and Wnt5a/Ror2/ABCA1/NF-kB trajectory

Atherosclerosis (AS) is a major cause of cardiovascular diseases that may lead to mortality. This study aimed to evaluate the therapeutic potential of tetrandrine in high cholesterol diet (HCD)-induced atherosclerosis, in rats, via modulation of miR-34a, as well as, Wnt5a/Ror2/ABCA1/NF-κB pathway and to compare its efficacy with atorvastatin. Induction of AS, in male rats, was done via IP administration of vitamin D3 (70 U/Kg for 3 days) together with HCD. At the end of the 9th week, rats were treated with atorvastatin at a dose of 20 mg/kg, and tetrandrine at different doses of (18.75, and 31.25 mg/kg) for 22 days. Serum inflammatory cytokines and lipid profile, liver oxidative stress parameters, and aortic tissue Wnt5a, Ror2, ABCA1, NF-κB, miR-34a levels were assessed in all experimental groups. Histopathological and Immunohistochemical assessments of aortic tissue sections were done. Results showed that tetrandrine treatment reverted the inflammatory and oxidative stress state together with reducing the serum lipids via modulating miR-34a, and Wnt5a/Ror2/ABCA1/NF-κB pathway. Moreover, it reverted the histopathological abnormalities observed in AS rats. Tetrandrine beneficial effects, in both doses, were comparable to that of atorvastatin, in most of the discussed parameters. These findings praise tetrandrine as a promising agent for management of atherosclerosis.

The expression of signal regulatory protein alpha (SIRPα) in periodontal cells and tissue.

Signal regulatory protein alpha (SIRPα) is mainly expressed by cells of myeloid origin. This membrane glycoprotein is shown to be involved in regulation of different inflammatory conditions, such as colitis and arthritis. However, SIRPα has not been investigated in relationship to periodontitis, an inflammatory condition affecting the tooth supporting tissues. We aim to investigate if resident cells in the periodontium express SIRPα and whether a possible expression is affected by inflammatory conditions. Primary human keratinocytes, fibroblasts, periodontal ligament cells, and osteoblasts were cultured with or without the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α) or interleukin-1-beta (IL-1β). All different periodontal cell types showed a basal mRNA expression of SIRPα. Pro-inflammatory cytokines induced a 2-3-fold significant increase in SIRPα expression in both cultured human gingival fibroblasts and osteoblasts but neither in keratinocytes nor in periodontal ligament cells. Tissue sections from human gingival tissue biopsies were histochemically stained for SIRPα. Epithelial keratinocytes and gingival fibroblasts stained positive in sections from periodontally healthy as well as in sections from periodontitis. In periodontitis sections, infiltrating leukocytes stained positive for SIRPα. We highlight our finding that oral keratinocytes, gingival fibroblasts, and periodontal ligament cells do express SIRPα, as this has not been presented before. The fact that inflammatory stimulation of gingival fibroblasts increased the expression of SIRPα, while an increased expression by gingival fibroblasts in periodontitis tissue in situ could not be detected, is indeed contradictory.

Taurine can play a positive role in growth, liver health and resistance to Aeromonas hydrophila of yellow catfish Pelteobagrus fulvidraco exposed to ammonia stress for a long time

Ammonia is an important environmental pollutant that is toxic to most aquatic animals, and nutrient manipulation is an effective regulatory pathway. To assess the influence of taurine supplementation on growth, liver detoxification and anti-bacterial infection of yellow catfish subjected to ammonia exposure for a long time, three experiment diets with different taurine levels (0.50, 1.00, and 2.00 %) were formulated and fed to fish (7.28 ± 0.05) g under three ammonia levels (0.00, 25.00 and 50.00 mg/L total ammonia nitrogen) for 56 days. The results showed that hepatic glycogen and whole-body protein contents were reduced, suggesting that energy deficiency caused by ammonia poisoning plays an important role in growth inhibition; elevated serum alanine aminotransferase and aspartate aminotransferase levels suggest liver injury, which may be caused by oxidative stress (decreased activities of superoxide dismutase and catalase, and expressions of SOD and CAT) and inflammation (increased contents of tumor necrosis factor α, interleukin 1 and interleukin 8, and expressions of TNF α, IL 1 and IL8), analysis of tissue sections further confirmed this finding. Furthermore, after injection with Aeromonas hydrophila, cumulative mortality increased proportionally with the rise in ammonia levels, the observed decrease in serum complements C3, C4, immunoglobulin M, and antibody titer contents emerged as a crucial factor contributing to the decreased disease resistance. The exogenous taurine was found to alleviate the negative impact of the above-mentioned ammonia poisoning. This study provides valuable insights into the mechanism of ammonia-induced aquatic toxicology in fish.

Full-length target sequences of GeoMx digital spatial profiling probes reveal that gene-promiscuity predicts probe sensitivity to EDTA tissue decalcification

GeoMx Digital Spatial Profiling (Nanostring) is a commercial spatial transcriptomics method to selectively analyze regions of interest within intact tissue sections. We show that decalcification with ethylene-diamine-tetra-acetic (EDTA) variably attenuates probe counts, while probes that are more resistant to this effect consequently appear overexpressed after quantile normalization. By determining the undisclosed full-length target sequences of probes used in the human whole transcriptome panel, hereby updating target transcripts and genes, we find that the gene-promiscuity of probes is an important factor that determines sensitivity to EDTA incubation.

The preventive effects of natural plant compound carvacrol against combined UVA and UVB-induced endoplasmic reticulum stress in skin damage of rats.

The skin is constantly exposed to a variety of environmental stressors, including ultraviolet (UV) radiation. Exposure of the skin to UV radiation causes a number of detrimental biological damages such as endoplasmic reticulum (ER) stress. The ER stress response is a cytoprotective mechanism that maintains homeostasis of the ER by increasing the capacity of the ER against the accumulation of unfolded proteins in the ER. Carvacrol (CRV) is a monoterpenoid phenol found in essential oils with antimicrobial and anti-inflammatory activities. We investigated for the first time in the literature the potential protective role of CRV against combined UVA and UVB-induced skin damage by targeting the ER stress pathway in a rat model. For this purpose, expressions of Grp78, Perk, Atf6, Ire-1, Chop, Xbp1, Casp12, elF2α, and Traf2 genes related to ER stress were analyzed by RT-PCR and protein expression levels of GRP78, ATF6, CHOP, and XBP1 were determined by ELISA assay in tissue sections taken from the back of the rats. As a result of analysis, it was seen that the expression levels of aforementioned ER stress genes increased significantly in the UVA + UVB irradiated group compared to the control group, while their expression levels decreased markedly by supplementation of CRV in UVA + UVB + CRV group. With regard to expressions of foregoing proteins, their levels escalated notably with UVA + UVB application and decreased markedly by CRV supplementation. In conclusion, present study revealed that CRV ameliorates UVA + UVB-induced ER stress via reducing the expression of mRNA as well as proteins involved in the unfolded protein response (UPR) pathway and inducing apoptosis as evidenced from high Caspase12 level.

Sonothrombolysis Using Microfluidically Produced Microbubbles in a Murine Model of Deep Vein Thrombosis.

The need for safe and effective methods to manage deep vein thrombosis (DVT), given the risks associated with anticoagulants and thrombolytic agents, motivated research into innovative approaches to resolve blood clots. In response to this challenge, sonothrombolysis is being explored as a technique that combines microbubbles, ultrasound, and thrombolytic agents to facilitate the aggressive dissolution of thrombi. Prior studies have indicated that relatively large microbubbles accelerate the dissolution process, either in an in vitro or an arterial model. However, sonothrombolysis using large microbubbles must be evaluated in venous thromboembolism diseases, where blood flow velocity is not comparable. In this study, the efficacy of sonothrombolysis was validated in a murine model of pre-existing DVT. During therapy, microfluidically produced microbubbles of 18 μm diameter and recombinant tissue plasminogen activator (rt-PA) were administered through a tail vein catheter for 30min, while ultrasound was applied to the abdominal region of the mice. Three-dimensional ultrasound scans were performed before and after therapy for quantification. The residual volume of the thrombi was 20% in animals post sonothrombolysis versus 52% without therapy ( ), indicating a significant reduction in DVT volume. Histological analysis of tissue sections confirmed a reduction in DVT volume post-therapy. Therefore, large microbubbles generated from a microfluidic device show promise in ultrasound-assisted therapy to address concerns related to venous thromboembolism.

SOX17 expression in mesonephric-like adenocarcinomas and mesonephric remnants/hyperplasia of the female genital tract: Expanding its utility as a Müllerian biomarker.

Recently, SOX17 has emerged as a promising biomarker for non-mucinous Müllerian (ovarian and endometrial) carcinomas, demonstrating increased specificity in comparison to PAX8 while maintaining similar sensitivity. However, expression of SOX17 in mesonephric-like adenocarcinoma (MLA), a carcinoma of the female genital tract with uncertain, but probably Müllerian histogenesis, remains unexplored. This study aims to address this gap. SOX17 immunohistochemistry was performed on whole tissue sections from 68 MLAs originating from the endometrium or ovary and seven cervical mesonephric carcinomas, as well as six mesonephric remnants/hyperplasias. Using a four-tiered scoring system based on distribution and intensity of staining, 68% of MLA displayed a negative/low (< 10%) SOX17 expression pattern, which contrasts with the high expression observed in most Müllerian carcinomas. However, 22% of MLA demonstrated high SOX17 expression, similar to other endometrial and ovarian carcinomas. Similarly, five of seven (72%) mesonephric carcinomas of the cervix were SOX17-negative, but two cases (28%) were positive. All mesonephric remnants/hyperplasias were SOX17 negative. The majority of MLA are negative or exhibit low SOX17 expression, in contrast to the diffuse and strong expression commonly seen in other types of Müllerian carcinoma. However, a subset of MLAs demonstrate high SOX17 expression. Therefore, absence of SOX17 staining is supportive for MLA when the differential includes another non-mucinous Müllerian carcinoma. SOX17 may also be useful for differentiating mesonephric remnants/hyperplasias from Müllerian malignancies and benign Müllerian glandular lesions.

First report of Diaporthe humulicola causing halo blight of hop (Humulus lupulus L.) in Prince Edward Island, Canada

This study documents the first report of halo blight of hop (Humulus lupulus L.), caused by the fungal pathogen Diaporthe humulicola, in Prince Edward Island (PE), Canada. This pathogen has been previously reported in Michigan, New York, Connecticut and Quebec. Disease symptoms were reported in hopyards in Harrington and Mount Albion, PE. Symptoms reported include both foliar lesions and severe necrosis of hop cones. Through a metagenomic approach using Nanopore sequencing of the full Internal Transcribed Spacer (ITS) region products, multiple fungal species were identified in infected hop cones. Many plant pathogens were present in the hop cones including species of Botrytis, Alternaria, Colletotrichum and Fusarium. Additionally, metagenomic sequencing identified reads that most closely matched D. humulicola. Symptomatic leaf tissue sections were cultured on amended PDA to isolate the pathogen, then colony PCR was conducted and ITS regions were sequenced, confirming the identity of D. humulicola. Pathogenicity was validated using Koch’s postulates on hop leaf disks, with lesion development occurring 7 days post-inoculation. Considering the humid conditions characteristic of eastern North American hop-growing zones, halo blight’s emergence in PE emphasizes the need for comprehensive monitoring and improved best management strategies to protect the PE hop industry.

Correlative Imaging for Comprehensive Molecular Mapping of Individual Cell Types in Biological Tissues.

Mass spectrometry imaging (MSI) is a powerful technique for label-free spatial mapping of multiple classes of biomolecules in tissue sections. However, differences in desorption and ionization efficiency of different classes of molecules make it challenging to simultaneously map biomolecules at each omics layer in the same tissue sample. Herein, we present a correlative imaging method using nanospray desorption electrospray ionization (nano-DESI) MSI, which enables the spatial mapping of lipids, metabolites, peptides, and proteins with cellular-level spatial resolution in a single tissue section. We demonstrate the molecular profiling of specific cell types and identify truncated peptides in mouse pancreatic tissue. Distinct chemical gradients of peptides and lipids extending from endocrine cells to exocrine cells indicate their different roles in endocrine-exocrine crosstalk and intracellular signaling. The results underscore the power of the developed imaging approach for spatial multi-omics analysis that provides deep insights into cellular diversity and the intricate molecular interactions that occur within heterogenous biological tissues.

Fetal Zika virus inoculation in macaques revealed control of the fetal viral load during pregnancy

BackgroundEarly pregnancy Zika virus (ZIKV) infection is associated with major brain damage in fetuses, leading to microcephaly in 0.6–5.0% of cases, but the underlying mechanisms remain largely unknown.MethodsTo understand the kinetics of ZIKV infection during fetal development in a nonhuman primate model, four cynomolgus macaque fetuses were exposed in utero through echo-guided intramuscular inoculation with 103 PFU of ZIKV at 70–80 days of gestation, 2 controls were mock inoculated. Clinical, immuno-virological and ultrasound imaging follow-ups of the mother/fetus pairs were performed until autopsy after cesarean section 1 or 2 months after exposure (n = 3 per group).ResultsZIKV was transmitted from the fetus to the mother and then replicate in the peripheral blood of the mother from week 1 to 4 postexposure. Infected fetal brains tended to be smaller than those of controls, but not the femur lengths. High level of viral RNA ws found after the first month in brain tissues and placenta. Thereafter, there was partial control of the virus in the fetus, resulting in a decreased number of infected tissue sections and a decreased viral load. Immune cellular and humoral responses were effectively induced.ConclusionsZIKV infection during the second trimester of gestation induces short-term brain injury, and although viral genomes persist in tissues, most of the virus is cleared before delivery.