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HPV-associated oral cavity squamous cell carcinoma (SCC) is not well-characterized in the literature, and also has a clinical significance that is poorly understood.We gathered a cohort of oral cavity (OC) SCC with nonkeratinizing morphology, either in the invasive or in situ carcinoma (or both), tested for p16 by immunohistochemistry and high risk HPV E6/E7 mRNA by RTPCR (reference standard for transcriptionally-active high risk HPV) and gathered detailed morphologic and clinicopathologic data.Thirteen patients from two institutions were proven to be HPV-associated by combined p16 and high risk HPV mRNA positivity. All 13 patients (100%) were males, all were heavy smokers (average 57 pack/year), and most were active drinkers (9/11 or 81.8%). All 13 (100%) involved the tongue and/or floor of mouth. All had nonkeratinizing features, but maturing squamous differentiation varied widely (0-90%; mean 37.3%). Nonkeratinizing areas had high N:C ratios and larger nests, frequently with pushing borders, and minimal (or no) stromal desmoplasia. The carcinoma in situ, when present, was Bowenoid/nonkeratinizing with cells with high N:C ratios, full thickness loss of maturation, and abundant apoptosis and mitosis. HPV was type 16 in 11 patients (84.6%) and type 33 in two (15.4%). Nine patients had treatment data available. These underwent primary surgical resection with tumors ranging from 1.6 to 5.2 cm. Most had bone invasion (6/9-66.7% were T4a tumors), and most (6/9-66.7%) had extensive SCC in situ with all 6 of these patients having final margins positive for in situ carcinoma.HPV-associated OCSCC is an uncommon entity that shows certain distinct clinical and pathologic features. Recognition of these features may help pathologic diagnosis and could potentially help guide clinical management.

PDF file - 384K, Detection of multiple HPV subtypes by degenerate PCR of genomic DNA (A) or by HPV-16 specific Southern blot (B).

XLSX file - 37K, Primer sequences used.

Purpose: To develop a clinically relevant model system to study head and neck squamous cell carcinoma (HNSCC), we have established and characterized a direct-from-patient tumorgraft model of human papillomavirus (HPV)–positive and HPV-negative cancers.Experimental Design: Patients with newly diagnosed or recurrent HNSCC were consented for donation of tumor specimens. Surgically obtained tissue was implanted subcutaneously into immunodeficient mice. During subsequent passages, both formalin-fixed/paraffin-embedded as well as flash-frozen tissues were harvested. Tumors were analyzed for a variety of relevant tumor markers. Tumor growth rates and response to radiation, cisplatin, or cetuximab were assessed and early passage cell strains were developed for rapid testing of drug sensitivity.Results: Tumorgrafts have been established in 22 of 26 patients to date. Significant diversity in tumorgraft tumor differentiation was observed with good agreement in degree of differentiation between patient tumor and tumorgraft (Kappa 0.72). Six tumorgrafts were HPV-positive on the basis of p16 staining. A strong inverse correlation between tumorgraft p16 and p53 or Rb was identified (Spearman correlations P = 0.085 and P = 0.002, respectively). Significant growth inhibition of representative tumorgrafts was shown with cisplatin, cetuximab, or radiation treatment delivered over a two-week period. Early passage cell strains showed high consistency in response to cancer therapy between tumorgraft and cell strain.Conclusions: We have established a robust human tumorgraft model system for investigating HPV-positive and HPV-negative HNSCC. These tumorgrafts show strong correlation with the original tumor specimens and provide a powerful resource for investigating mechanisms of therapeutic response as well as preclinical testing. Clin Cancer Res; 19(4); 855–64. ©2012 AACR.

XLSX - 49K, Antibodies, sources, and catalog numbers used throughout manuscript.

XLSX - 46K, Table of Primer sequences used and probes.

XLSX - 35K, Sources and culture conditions for cell lines.

Female reproductive performance is essential for economic profitability of swine operations, and sex steroid hormones, such as progesterone (P4), are necessary for proper reproductive function. Recent research targeting the reproductive microbiome in beef cattle found that as P4 concentrations fluctuate during the estrous cycle, bacterial communities in reproductive tissues also change. However, characterizing the relationship between P4 concentrations and bacterial communities within the reproductive tract has yet to be fully determined in swine. Therefore, the objective of this project was to correlate the abundance of reproductive tract bacterial communities with plasma P4 concentrations from pregnant gilts. Duroc X Landrace X Yorkshire gilts (n = 25), free of any physical or health related issues, were euthanized and hysterectomized on days (d) 11 (n = 7), 15 (n = 6), 60 (n = 6) or 90 (n = 6) of gestation. For microbiome evaluation, sterile swabs were individually rotated 8 times on each tissue and immediately placed in sterile microcentrifuge tubes for storage (-80°C) until sequencing. Allantoic and amniotic fluid collected on d 60 and d 90 of gestation was placed immediately in a sterile 10 mL conical tube and stored (-80°C) for subsequent analyses. Blood was collected concurrently, centrifuged at 2,500 x g for 25 min at 4°C, and the resulting plasma stored (-20°C). Concentrations of P4 in plasma were quantified using a double-antibody RIA kit. Bacterial sequencing of the V4 hypervariable region of 16S rRNA gene was completed by FERA Diagnostics and Biologicals Corp. The data were analyzed statistically using PROC CORR in SAS 9.4. In the cervix, the relative abundance of the phylum Firmicutes was positively correlated with P4 concentrations (r = 0.45, P = 0.03), and within this phylum, genus Lactobacillus was positively correlated with P4 concentrations (r = 0.54, P = 0.01). Phylum Bacteroidetes was negatively correlated with P4 concentrations (r = 0.45, P = 0.03), and within this phylum, genera Prevotella and Bacteroides were also negatively correlated with P4 concentrations (r = -0.49, P = 0.01; r = -0.05, P = 0.01, respectively). In the uterus, plasma P4 tended to be negatively correlated with phylum Proteobacteria (r = -0.37; P = 0.07) yet positively correlated with Phylum Firmicutes (r = 0.40; P = 0.05). Within Firmicutes, genus Streptococcus in the uterus was positively correlated with P4 concentrations (r = 0.50, P = 0.01), and the relative abundances of genus Lactobacillus in both the uterus and allantoic fluid were positively correlated with concentrations of P4 in plasma (r = 0.43, P = 0.04; r = 0.70, P = 0.02, respectively). There were no significant correlations between concentrations of P4 in plasma and abundances of bacteria within the vagina or amniotic fluid. These results indicate that shifts plasma P4 influence microbiomes in reproductive tissues of gilts during gestation. Further investigation is necessary to understand the interplay between P4 and microbiome fluctuations within reproductive tissues in swine.

Female sex steroid hormones have been shown to interact with cytokines within the immune system to help influence fertility. Previous research has indicated that shifts in commensal bacteria are associated with compromised reproductive performance during the postpartum period, yet evaluation of how the female sex hormone progesterone (P4) influences these microbial shifts is not fully understood. The objectives of this study were to determine 1) the relationship between P4 concentrations and cytokine concentrations, and 2) the relationship between P4 concentrations and the reproductive microbiome. Twenty Bos indicus-influenced beef cows (~60 days postpartum and free of health issues) were weighed, body condition scored and then subjected to the Bos indicus prostaglandin 5 day (d) + CIDR protocol beginning on d -8 and timed artificially inseminated on d 0. Blood samples were collected on d-3, -1, 0, and 28. Cytokines and reproductive microbiome were evaluated using vaginal and uterine flushes collected on d -3 and d-1. Experimental groups were assigned by d 28 pregnancy status, determined by transrectal ultrasonography, resulting in Open (n = 13) and Pregnant (n = 7) females. Bacterial community analyses were conducted targeting the V4 hypervariable region of the 16S rRNA gene. Plasma P4 concentrations on d -3 and d -1 were quantified per manufacturers’ instructions using a double-antibody RIA kit. Cytokine analyses of plasma and flushes were conducted using the RayBiotech Quantibody Bovine Cytokine Array Q1 kit per the manufacturer’s instructions. Statistical analyses were conducted using the GLM procedure and CORR procedure in SAS. Plasma P4 concentrations did not differ by pregnancy status (P > 0.05) but did decrease from d -3 to d -1 (0.89 vs. 0.08 ± 0.23 ng/mL, respectively; P = 0.02). There were no significant correlations between plasma P4 and plasma cytokines (P > 0.05). Interferon-alpha (IFNα) and P4 concentrations were positively correlated in Open and Pregnant uterine flushes (r = 0.41; P = 0.01). Concentrations of P4 interleukin (IL)-1F5 were negatively correlated in Open and Pregnant vaginal flushes (r = -0.32; P = 0.04). The relative abundance of genus Methanobrevibacter in uterine flushes was positively correlated with P4 concentrations in both experimental groups (r = 0.35; P = 0.02). Within vaginal flushes, there was a tendency for a negative correlation between the relative abundance of genus Bacteroides and P4 concentrations (r = -0.31; P = 0.05). Alternatively, there is a positive correlation between P4 concentration and relative abundance of genus Streptococcus (r = 0.47; P = 0.01) and genus Methanobrevibacter (r = 0.37; P = 0.02) in vaginal flushes. These results suggest there may be relationships between several cytokines and circulating P4 concentrations; therefore, potentially suggesting immunomodulatory activities of P4. Overall, this research alludes to P4 having a vital role in reproductive microbial shifts and associated immune responses. Further investigation on the local effects of P4 within the reproductive tract on immune responses and bacterial shifts could provide greater insight into mechanisms to improve fertility in beef cattle.

The differences between the microbial communities in reproductive tissues, and amniotic and allantoic fluids of cattle have previously been investigated. Our objective was to provide insight into microbial differences across reproductive tissues in gilts during mid- and late-gestation. Specifically, we characterized the microbiomes of the vagina, cervix, endometrium, chorion, and allantoic and amniotic fluids. Duroc x Landrace x Yorkshire gilts (n = 12) free of physical, health or reproductive-related issues were euthanized and hysterectomized at either mid-gestation (d 60, n = 6) or late-gestation (d 90, n = 6). A sterile swab was rotated 8 times to collect samples from the mucosal surface of individual tissues and immediately placed in microcentrifuge tubes for storage (-80°C) until sequencing. To sample the endometrium and cervix, a 1 cm incision was made to expose the endometrium and interdigitating prominences, whereas for vaginal sampling, sterile swabs were inserted 6 inches past the vulva and then rotated. For collection of samples of chorion, an incision was made in the endometrium, exposing the chorioallantois to allow sampling of both the chorion-endometrial and chorion-allantois sides. For allantoic and amniotic fluids, 5 mL samples were collected and placed immediately in a sterile 10 mL conical tube and stored (-80°C) for subsequent analyses. Bacterial DNA was extracted and genome sequencing targeting the V4 hypervariable region of the 16S rRNA gene was conducted by FERA Diagnostics and Biologicals Corp. Data were analyzed statistically using the GLM procedure in SAS. Across all samples, abundances of the phylum Firmicutes was affected by stage of pregnancy with greater relative abundance at d 90 compared with d 60 (52.23 ± 2.85% vs 42.60 ± 3.14%, respectively; P = 0.03). Within this phylum and across all samples, the relative abundance of the genus Lactobacillus differed due to stage of pregnancy with a greater relative abundance at d 90 compared with d 60 (19.33 ± 2.37% vs 12.14 ± 2.61%, respectively; P = 0.05). Lactobacillus was most abundant in allantoic fluid when compared with all other samples (P < 0.01). The relative abundance of the genus Streptococcus, within the phylum Firmicutes, was less in amniotic and allantoic fluid compared with the cervix and vagina (P < 0.01). The phylum Bacteroidetes tended to differ by sample type (P = 0.06) with amniotic fluid having the lowest relative abundance. Within this phylum, the relative abundance of the genus Prevotella was also least in amniotic fluid (P < 0.01). These results suggest that microbial communities of reproductive tissues and fluids differ between mid- and late-gestation in pregnant gilts. Additional research on reproductive microbiomes in swine, and other livestock species, is required to develop methods to modulate bacteria and improve reproductive performance and successful outcomes of pregnancy.

The pioneer microbiome is the initial and foundational colonization of microbiological organisms within the neonatal animal and has lifelong implications for health and performance. The objective of this project was to quantify various maternal and environmental sources contributing to the pioneer microbiome of the piglet. Five pregnant gilts farrowed at the Texas A&M University Swine Center in April 2022. Litters were a Duroc x Landrace x Yorkshire composite and averaged 13.4 total number born and 11.4 number weaned. At birth, all piglets were individually identified and weighed and 5 piglets per litter weighing ≥1,200 grams were selected for microbiome analysis. Sterile swabs were used to collect, in duplicate, samples from the farrowing crate before gilt introduction (EMPTYCRATE) and after gilts were moved in (FULLCRATE), from the birth canal during farrowing (BIRTHCANAL), and from the rectum of the piglet on days 0 (prior to suckling), 3, and 10 post-farrowing and at weaning (21.6 ±.0 days post-farrowing). Colostrum was collected from each gilt during farrowing from a representative sample of teats into a single sterile collection cup (COLOSTRUM). Swabs and colostrum were stored in sterile microcentrifuge tubes at -80°C until sequencing. Bacterial DNA was extracted, and sequencing targeted the V4 hypervariable region of the6S rRNA gene. Statistical analyses were conducted using PROC GLM and PROC REG in SAS 9.4. The phyla Firmicutes and Proteobacteria were not different between maternal (BIRTHCANAL, COLOSTRUM) and environmental (EMPTYCRATE, FULLCRATE) sources (Table 1). The piglet microbiome shifted from birth to weaning. The relative abundance of phylum Firmicutes was less on day 3 compared with day 0 and at weaning but did not differ from day0 (Table 1). Within the phylum Firmicutes, Lactobacillus and Clostridium genera were greater on day 3 compared with day 0, 10, and at weaning (Table 1). The relative abundance of phylum Proteobacteria, and the relative abundance of genus Escherichia within phylum Proteobacteria, were greater on day 3 compared with day 0, 10, and at weaning (Table 1). Multiple regression analyses indicated that BIRTHCANAL explained 51.6% of the variation observed in piglet day 0 microbiome (P < 0.0001), and 6.5% of the variation in the piglet day 10 microbiome (P = 0.01). The piglet day 0 (pre-suckle) microbiome explained 10.0% of the variation observed in day 3 microbiome and 15.6% of the day 10 microbiome (P < 0.05). Finally, day 10 microbiome explained 58.6% of the variation observed in the piglet microbiome at weaning (P < 0.0001). The microbiome of COLOSTRUM and the farrowing crate did not impact piglet microbiome (P > 0.10). Results indicate that the piglet pioneer microbiome is largely influenced by the microbiome of the birth canal and that the microbiome of the animal at weaning may be largely established by 10 days of age.

Status of bacterial diversity within the male seminal microbiome has previously been linked to fertility status in mammals. However, minimal research has focused on the reproductive microbiome in boar semen. The objective of this study was to analyze the diversity of bacterial communities within fresh extended boar semen samples 1) between studs, 2) over 5 consecutive days, and 3) between pooled and single-sire doses. Eight single-sire (n = 4 per stud) and eight pooled (n = 4 per stud) fresh extended semen doses (80 mL) were obtained from two boar studs (A vs. B). Pooled doses were the composite of boars in single-sire doses. Doses were subsampled (16 mL) for 5 days post-collection. Negative controls remained unopened until the last day. Each day, 5 mL of each subsample were flash frozen and stored at -80°C for bacterial analysis targeting the V4 hypervariable region of the 16S rRNA gene for Illumina sequencing. Quality of all sequences was determined by FastQC before being processed for taxonomic analysis using the qiime2 pipeline. Alpha diversity (difference within sample metric) was analyzed using Kruskal-Wallis test, whereas PERMANOVA was performed for beta diversity analysis (differences between sample metric). According to alpha diversity metrics, Chao1 (P < 0.01) and Shannon’s index (P < 0.01) indicated contrasts in diversity among all days and the negative controls. This was further confirmed with a pairwise comparison between individual groups, in which day 1 exhibited lower alpha diversity when compared with days 2, 3, 4, and 5 using Chao1 (P < 0.01) and Shannon’s index (P < 0.01). Interestingly, reduced diversity was observed for day 1 compared with the negative controls with Chao1 (57.4 ± 6.6 vs. 132.4 ± 22.1, P < 0.01, Q < 0.01; respectively) and Shannon’s index (4.8 ± 0.2 vs. 6.1 ± 0.3, P < 0.01, Q < 0.01; respectively). No differences between boar stud or sire-type were detected for alpha diversity metrics. For beta diversity analyses, both PCoA plots and PERMANOVA for weighted unifrac metric demonstrated no significant clustering between sampling days. However, distinctions between all days and negative controls were identified using PERMANOVA for unweighted unifrac metric. Specifically, differences were discovered between day 1 and days 3, 4, and 5 (P < 0.01, Q < 0.01). PERMANOVA for both weighted (P < 0.01) and unweighted (P < 0.01) unifrac metrics revealed distinct clustering by boar stud. However, there were no differences in beta diversity between pooled and single-sire samples for both weighted and unweighted unifrac metrics. Based on these results, bacterial diversity within a semen sample may be altered over time and could occur between boar stud operations. Additional studies analyzing the reproductive microbiome of semen could help improve the characterization of fertility status in boars.

Evaluation of bone pathology within the head and neck region, particularly the gnathic bonesis is complex, demonstrating unique pathologic processes. In part, this variation is due to odontogenesis and the embryological cells that may be involved, which can contribute to disease development and histologic variability. As with any boney pathosis, the key is to have clinical correlation, particularly with radiographic imaging prior to establishing a definitive diagnosis. This review will cover those entities that have a predilection for the pediatric population, and while it is not all inclusive, it should serve as a foundation for the pathologist who is evaluating bony lesions involving the craniofacial skeleton.

Translational animal models for oral mucositis (OM) are necessary to simulate and assess the bioclinical effects and response in humans. These models should simulate high levels of radiation exposure that leads to oxidative stress and inflammatory‐initiated tissue changes. Hamster models have been extensively studied to observe pathological effects of radiation exposure and help in the development of effective treatments. To successfully evaluate the potential for treatment regimens with consistency and relevance, a radiation‐induced OM hamster model was developed using a clinical linear accelerator utilized by cancer patients daily. The dose exposure to the isolated, everted cheek pouch of a hamster, as well as the progression of injury, pro‐inflammatory marker, histological, and elasticity analyses of the buccal pouch were conducted to verify replicability and reproducibility of the injury model. The findings from this model demonstrated its ability to consistently induce injury and resolution over 28 days using an acute dose of 60 Gy. This model was developed to enhance clinical relevance when evaluating potential efficacious treatments and can now be utilized in efficacy studies to better evaluate developed therapeutics in a preclinical model that is easy to translate to clinical studies..

BACKGROUND: Syphilis is a sexually-transmitted infectious disease caused by Treponema pallidum. Cases of primary and secondary syphilis are on the rise in the United States, with a 14.4% increase in new cases noted from 2017 to 2018 and an escalation of 71% between the years 2014 and 2018. Fulfilling its nickname of “the great imitator,” oral manifestations of syphilis may mimic a variety of infectious, neoplastic, or immune-mediated processes, both clinically and histopathologically. This large spectrum of appearances can create a diagnostic challenge to the clinician and/or pathologist, leading to delay in diagnosis or misdiagnosis. METHODS: A database of oral syphilis cases was created from archives at the University of Kentucky, University of Pittsburgh, LIJMC, Columbia University MC, and University of Tennessee. The age, sex, race, location, duration, and clinical description were recorded. Cases without positive reaction upon immunohistochemistry or serologic tests were excluded. RESULTS: We identified 19 new cases of oral syphilis (17 males, one female, and one case unknown sex) and described the clinical and histopathological features of this re-emerging and potentially fatal disease. All cases demonstrated dense lymphoplasmacytic inflammation, often with inflammatory exocytosis or ulceration at the surface, and perivascular inflammation. CONCLUSIONS: Early recognition of the histopathologic and clinical manifestations of oral syphilis is imperative for prompt diagnosis, improved patient outcomes, and disease prevention.

Shifts in commensal bacteria (i.e., Firmicutes) in the human microbiome are associated with compromised fertility. The objectives of this study were to characterize 1) the reproductive microbiome and 2) the cytokine concentrations prior to artificial insemination (AI) in vaginal and uterine flushes from postpartum beef cows. Twenty Bos indicus-influenced beef cows (~60 days postpartum and free of health issues) were weighed, body condition scored (BCS) then subjected to the Bee Synch II synchronization protocol on d-8 and timed AI on d0. Blood samples were collected on d-3, -1, 0, and 28. Vaginal and uterine flushes were collected on d-3 and d-1. On d28, pregnancy was determined by transrectal ultrasonography (Open, n=13 and Pregnant, n=7). Bacterial community analyses were conducted targeting the V4 hypervariable region of the 16S rRNA gene. Cytokine analyses were conducted using the RayBiotech Quantibody® Bovine Cytokine Array Q1 kit per manufacturer's instructions. Weight, BCS, and vaginal bacterial communities by phyla were not different (P >0.05). In open cows, the uterine relative abundance of Firmicutes decreased between d-3 and -1 (60.4% ± 0.9% vs. 48.5% ± 3.2%, respectively; P< 0.05). In pregnant cows, the uterine relative abundance of Tenericutes significantly increased between d-3 and -1 (1.0% ± 0.1% vs. 7.6% ± 4.1%, respectively; P< 0.05). Additionally, the genus Ureaplasma tended to increase in relative abundance from d-3 to -1 (0.1% ± 0.1% vs. 7.3% ± 4.9%, respectively; P=0.06). Uterine concentrations of Interferon (IFN)-γ, Interleukin (IL)-1α and IL-21 were greater in open females compared to pregnant. Vaginal concentrations for IFN-α and chemokine ligand (CXCL)-10 were greater in open females compared to pregnant. Regardless of treatment, IL-13 increased from d-3 to d-1. These results suggest a distinct difference in the uterine microbiome and cytokine profiles prior to AI for resulting pregnant and open cows.

Bacterial contamination of non-frozen extended boar semen has been associated with a decrease in sperm viability and longevity. The objective of this study was to analyze the microbiome of extended boar semen and sperm quality parameters 1) between studs 2) over time and 3) between pooled and single-sire doses. Sixteen pooled (n=4 per stud) and single-sire (n=4 per stud) boar semen doses (80mL) were obtained from two studs (A vs. B). Pooled doses were the composite of boars in single-sire doses. Doses were stored at 16°C and subsampled 16mL daily for five days post-collection. Each day, 5mL were flash frozen and stored at -80°C for bacterial analysis targeting the V4 hypervariable region of the 16S rRNA gene. The remaining 11mL were used to assess sperm progressive motility and agglutination (0: no adhesion to 3: >50% adhesion) by two evaluators with 10 visual estimates per sample. Negative controls of each pooled (n=2 per stud) and single-sire (n=4 per stud) dose remained unopened until day 5. Statistical analyses were conducted using GLM procedure in SAS. Sperm agglutination was greater in stud A than B (1.6 vs. 1.0 ± 0.1, respectively; P< 0.01). Sperm motility tended (P=0.09) to be greater in stud B than A doses (67.4% vs. 61.5% ± 0.02%, respectively), and decreased over time (P< 0.01). Stud B had a greater relative abundance of Proteobacteria (60.0% vs. 47.2% ± 1.5%, respectively; P< 0.01) and lower abundance of Firmicutes (22.5% vs. 31.9% ± 1.4%, respectively; P< 0.01) compared to stud A. Bacterial load did not differ by stud or dose type but increased from day 1 to 3 (P< 0.01). Bacterial load between days 3, 4, and 5 post-collection was not different from the negative control. Results indicate differences in the microbiome between boar studs and its influence on semen over time.

Cytokines have a vital role in reproductive immune environment during the postpartum period. Previous data evaluated uterine and vaginal cytokine concentrations prior to insemination; however, was unable to correlate these data with plasma cytokine concentrations. The objective of this study was to determine relationships between uterine, vaginal, and plasma pro- and anti-inflammatory cytokine concentrations in postpartum beef cows prior to timed artificial insemination (TAI). Bos indicus-influenced beef cows (n=20) free of any physical, health or reproductive-related issues were subjected to the Bee Synch II protocol 8 days prior (d-8) to TAI (d0). Uterine and vaginal flushes and blood were collected on d-3 and d-1. Pregnancy was determined by transrectal ultrasonography on d28 (Pregnant, n=7; Open, n=13). Using the RayBiotech Quantibody Bovine Cytokine Array 1 the following cytokines concentrations were determined: interferon (IFN)-α, IFN-γ, interleukin (IL)-13, IL-1α, IL-1-F5, IL-21, tumor necrosis factor (TNF)-α, chemokine ligand (CXCL)-9, CXCL-10, and chemokine ligand 4 (CCL4). Concentration data were analyzed using PROC GLM and correlations using Pearson correlation in SAS. For plasma samples, 8 pro-inflammatory cytokines (IFN-α, IFN-γ, IL-13, IL-1α, IL-1-F5, IL-21, CXCL-9, and TNF-α) had greater (P< 0.05) concentrations in open cows compared with pregnant cows. For uterine flushes, 3 pro-inflammatory cytokines (IFN-γ, IL-1α, and IL-21) had greater (P< 0.05) concentrations in open cows compared with pregnant cows. For vaginal flushes, IFN-α concentrations were greater (P< 0.05) in open cows, while CXCL-10 concentrations were greater in pregnant cows. There were no significant correlations between plasma and uterine or vaginal samples (P>0.05). Interestingly, for open cows there were correlations for IFN-α (r=0.46; P=0.02), IFN-γ (r=0.58; P=0.002), and IL-21 (r=0.54; P=0.004) between uterine and vaginal samples; however, no correlations were observed for pregnant cows. These results suggest a greater abundance of pro-inflammatory cytokines within the uterus, vagina, and in peripheral circulation for resulting open cows prior to TAI.

Dysregulation of innate immune signaling pathways is implicated in various hematologic malignancies. However, these pathways have not been systematically examined in acute myeloid leukemia (AML). We report that AML hematopoietic stem and progenitor cells (HSPCs) exhibit a high frequency of dysregulated innate immune-related and inflammatory pathways, referred to as oncogenic immune signaling states. Through gene expression analyses and functional studies in human AML cell lines and patient-derived samples, we found that the ubiquitin-conjugating enzyme UBE2N is required for leukemic cell function in vitro and in vivo by maintaining oncogenic immune signaling states. It is known that the enzyme function of UBE2N can be inhibited by interfering with thioester formation between ubiquitin and the active site. We performed in silico structure-based and cellular-based screens and identified two related small-molecule inhibitors UC-764864/65 that targeted UBE2N at its active site. Using these small-molecule inhibitors as chemical probes, we further revealed the therapeutic efficacy of interfering with UBE2N function. This resulted in the blocking of ubiquitination of innate immune- and inflammatory-related substrates in human AML cell lines. Inhibition of UBE2N function disrupted oncogenic immune signaling by promoting cell death of leukemic HSPCs while sparing normal HSPCs in vitro. Moreover, baseline oncogenic immune signaling states in leukemic cells derived from discrete subsets of patients with AML exhibited a selective dependency on UBE2N function in vitro and in vivo. Our study reveals that interfering with UBE2N abrogates leukemic HSPC function and underscores the dependency of AML cells on UBE2N-dependent oncogenic immune signaling states.

Tropical cyclones (TCs) moving into the midlatitudes can produce extreme precipitation, as was the case with Hurricane Irene (2011). Despite the high-impact nature of these events, relatively few studies have explored the sensitivity of TC precipitation forecasts to model initial conditions. Here, the physical processes that modulate precipitation forecasts over the Northeast United States during Irene are investigated using an 80-member 0.5° Global Forecasting System (GFS) ensemble. The members that forecast the highest total precipitation over the Catskill Mountains in New York (i.e., wet members) are compared with the members that predicted the least precipitation (i.e., dry members). Results indicate that the amount of rainfall is tied to storm track, with the wetter members forecast to move farther west than the dry members. This variability in storm track appears to be associated with variability in analyzed upper-tropospheric potential vorticity (PV), such that the wetter members feature greater cyclonic PV southwest of Irene when Irene is off the Carolina coast. By contrast, the wetter members of a 3-km Weather Research and Forecasting (WRF) Model ensemble, initialized from the same GFS ensemble forecasts, show little sensitivity to track. Instead, the wetter members are characterized by stronger lower-tropospheric winds perpendicular to the eastern face of the Catskills, allowing maximum upslope forcing and horizontal moisture flux convergence during the period of heaviest rainfall. The drier members, on the other hand, have the greatest quasigeostrophic forcing for ascent, implying that the members’ differences in mesoscale topographic forcing are the dominant influence on rainfall rate.

Despite evidence of chronic inflammation in myelodysplastic syndrome (MDS) and cell-intrinsic dysregulation of Toll-like receptor (TLR) signaling in MDS hematopoietic stem and progenitor cells (HSPCs), the mechanisms responsible for the competitive advantage of MDS HSPCs in an inflammatory milieu over normal HSPCs remain poorly defined. Here, we found that chronic inflammation was a determinant for the competitive advantage of MDS HSPCs and for disease progression. The cell-intrinsic response of MDS HSPCs, which involves signaling through the noncanonical NF-κB pathway, protected these cells from chronic inflammation as compared to normal HSPCs. In response to inflammation, MDS HSPCs switched from canonical to noncanonical NF-κB signaling, a process that was dependent on TLR-TRAF6-mediated activation of A20. The competitive advantage of TLR-TRAF6-primed HSPCs could be restored by deletion of A20 or inhibition of the noncanonical NF-κB pathway. These findings uncover the mechanistic basis for the clonal dominance of MDS HSPCs and indicate that interfering with noncanonical NF-κB signaling could prevent MDS progression.

Epstein-Barr virus (EBV)-positive extranodal marginal zone lymphomas of mucosa-associated lymphoid tissue (MALT lymphomas) were initially described in solid organ transplant recipients, and, more recently, in other immunodeficiency settings. The overall prevalence of EBV-positive MALT lymphomas has not been established, and little is known with respect to their genomic characteristics. Eight EBV-positive MALT lymphomas were identified, including 1 case found after screening a series of 88 consecutive MALT lymphomas with EBER in situ hybridization (1%). The genomic landscape was assessed in 7 of the 8 cases with a targeted high throughput sequencing panel and array comparative genomic hybridization. Results were compared to published data for MALT lymphomas. Of the 8 cases, 6 occurred post-transplant, 1 in the setting of primary immunodeficiency, and 1 case was age-related. Single pathogenic/likely pathogenic mutations were identified in 4 of 7 cases, including mutations in IRF8, BRAF, TNFAIP3, and SMARCA4. Other than TNFAIP3, these genes are mutated in3% of EBV-negative MALT lymphomas. Copy number abnormalities were identified in 6 of 7 cases with a median of 6 gains and 2 losses per case, including 4 cases with gains in regions encompassing several IRF family or interacting genes (IRF2BP2, IRF2, and IRF4). There was no evidence of trisomies of chromosomes 3 or 18. In summary, EBV-positive MALT lymphomas are rare and, like other MALT lymphomas, are usually genetically non-complex. Conversely, while EBV-negative MALT lymphomas typically show mutational abnormalities in the NF-κB pathway, other than the 1 TNFAIP3-mutated case, no other NF-κB pathway mutations were identified in the EBV-positive cases. EBV-positive MALT lymphomas often have either mutations or copy number abnormalities in IRF family or interacting genes, suggesting that this pathway may play a role in these lymphomas.

We are building an experiment to search for dark matter in the form of dark photons in the nano- to milli-eV mass range. This experiment is the electromagnetic dual of magnetic detector dark radio experiments. It is also a frequency-time dual experiment in two ways: We search for a high-Q signal in wide-band data rather than tuning a high-$Q$ resonator, and we measure electric rather than magnetic fields. In this paper we describe a pilot experiment using room temperature electronics which demonstrates feasibility and sets useful limits to the kinetic coupling $\epsilon \sim 10^{-12}$ over 50--300 MHz. With a factor of 2000 increase in real-time spectral coverage, and lower system noise temperature, it will soon be possible to search a wide range of masses at 100 times this sensitivity. We describe the planned experiment in two phases: Phase-I will implement a wide band, 5-million channel, real-time FFT processor over the 30--300 MHz range with a back-end time-domain optimal filter to search for the predicted $Q\sim 10^6$ line using low-noise amplifiers. We have completed spot frequency calibrations using a biconical dipole antenna in a shielded room that extrapolate to a $5 \sigma$ limit of $\epsilon\sim 10^{-13}$ for the coupling from the dark field, per month of integration. Phase-II will extend the search to 20 GHz using cryogenic preamplifiers and new antennas., Comment: 11 pages, 13 figures. Updated to published version. Corrected minor error in Fig 12 x-axis; results unchanged