Geographic variation of mutagenic exposures in kidney cancer genomes.

International differences in the incidence of many cancer types indicate the existence of carcinogen exposures that have not yet been identified by conventional epidemiology make a substantial contribution to cancer burden1. In clear cell renal cell carcinoma, obesity, hypertension and tobacco smoking are risk factors, but they do not explain the geographical variation in its incidence2. Underlying causes can be inferred by sequencing the genomes of cancers from populations with different incidence rates and detecting differences in patterns of somatic mutations. Here we sequenced 962 clear cell renal cell carcinomas from 11 countries with varying incidence. The somatic mutation profiles differed between countries. In Romania, Serbia and Thailand, mutational signatures characteristic of aristolochic acid compounds were present in most cases, but these were rare elsewhere. In Japan, a mutational signature of unknown cause was found in more than 70% of cases but in less than 2% elsewhere. A further mutational signature of unknown cause was ubiquitous but exhibited higher mutation loads in countries with higher incidence rates of kidney cancer. Known signatures of tobacco smoking correlated with tobacco consumption, but no signature was associated with obesity or hypertension, suggesting that non-mutagenic mechanisms of action underlie these risk factors. The results of this study indicate the existence of multiple, geographically variable, mutagenic exposures that potentially affect tens of millions of people and illustrate the opportunities for new insights into cancer causation through large-scale global cancer genomics.

The Complexity of Tobacco Smoke-Induced Mutagenesis in Head and Neck Cancer.

Tobacco smoke, alone or combined with alcohol, is the predominant cause of head and neck cancer (HNC). Here, we further explore how tobacco exposure contributes to cancer development by mutational signature analysis of 265 whole-genome sequenced HNC from eight countries. Six tobacco-associated mutational signatures were detected, including some not previously reported. Differences in HNC incidence between countries corresponded with differences in mutation burdens of tobacco-associated signatures, consistent with the dominant role of tobacco in HNC causation. Differences were found in the burden of tobacco-associated signatures between anatomical subsites, suggesting that tissue-specific factors modulate mutagenesis. We identified an association between tobacco smoking and three additional alcohol-related signatures indicating synergism between the two exposures. Tobacco smoking was associated with differences in the mutational spectra and repertoire of driver mutations in cancer genes, and in patterns of copy number change. Together, the results demonstrate the multiple pathways by which tobacco smoke can influence the evolution of cancer cell clones.

Assigning mutational signatures to individual samples and individual somatic mutations with SigProfilerAssignment.

MOTIVATION: Analysis of mutational signatures is a powerful approach for understanding the mutagenic processes that have shaped the evolution of a cancer genome. To evaluate the mutational signatures operative in a cancer genome, one first needs to quantify their activities by estimating the number of mutations imprinted by each signature. RESULTS: Here we present SigProfilerAssignment, a desktop and an online computational framework for assigning all types of mutational signatures to individual samples. SigProfilerAssignment is the first tool that allows both analysis of copy-number signatures and probabilistic assignment of signatures to individual somatic mutations. As its computational engine, the tool uses a custom implementation of the forward stagewise algorithm for sparse regression and nonnegative least squares for numerical optimization. Analysis of 2700 synthetic cancer genomes with and without noise demonstrates that SigProfilerAssignment outperforms four commonly used approaches for assigning mutational signatures. AVAILABILITY AND IMPLEMENTATION: SigProfilerAssignment is available under the BSD 2-clause license at https://github.com/AlexandrovLab/SigProfilerAssignment with a web implementation at https://cancer.sanger.ac.uk/signatures/assignment/.

Visions of recovery: a cross-diagnostic examination of eating disorder pro-recovery communities on TikTok.

Individuals seeking support or inspiration for eating disorder recovery may turn to pro-recovery content on social media sites such as TikTok. While research has thus far treated pro-recovery social media as a fairly homogeneous space, many pro-recovery hashtags single out particular eating disorder diagnoses. This exploratory study used codebook thematic analysis of 241 popular pro-recovery videos on TikTok to compare the presentation of eating disorders and eating disorder recovery across five different diagnosis-specific hashtags: #anarecovery, #arfidrecovery, #bedrecovery, #miarecovery, and #orthorexiarecovery. These hashtags refer to the following eating disorder diagnoses respectively: anorexia nervosa, avoidant restrictive food intake disorder, binge eating disorder, bulimia nervosa, and orthorexia nervosa. Our analysis generated the following qualitative themes across the entire dataset: (1) centrality of food to eating disorders and recovery, (2) what eating disorders look and feel like, (3) recovery as a process, (4) getting and giving help, and (5) negotiating diet culture in recovery. To supplement our qualitative findings and facilitate cross-diagnostic comparisons, we also conducted one-way ANOVAs and chi-square tests to probe for statistically significant differences in audience engagement and code prevalence across the different hashtags. Our results indicate that there are clear differences in how recovery is envisioned on TikTok based on which diagnostic hashtags are employed. Such variations in how different eating disorders are imagined on popular social media demand further investigation and clinical consideration.

Hashtags related to eating disorder recovery on TikTok often name particular eating disorder diagnoses. This study compared five of these diagnosis-specific hashtags: #anarecovery, #arfidrecovery, #bedrecovery, #miarecovery, and #orthorexiarecovery. We found some similarities among these hashtags such as the centrality of food and eating in the posts and the emphasis on recovery as a process. However, we also found significant differences between the hashtags. For example, while diet culture promotion was a key aspect of many #bedrecovery posts, #orthorexiarecovery posts tended to focus instead on critiques of diet culture. Levels of user engagement also varied across the five hashtags. Notably, #anarecovery posts received the most likes. This study points to the existence of subcommunities within pro-recovery social media and suggests that way eating disorder recovery is portrayed online differs across diagnostic labels.

Assigning mutational signatures to individual samples and individual somatic mutations with SigProfilerAssignment.

Analysis of mutational signatures is a powerful approach for understanding the mutagenic processes that have shaped the evolution of a cancer genome. Here we present SigProfilerAssignment, a desktop and an online computational framework for assigning all types of mutational signatures to individual samples. SigProfilerAssignment is the first tool that allows both analysis of copy-number signatures and probabilistic assignment of signatures to individual somatic mutations. As its computational engine, the tool uses a custom implementation of the forward stagewise algorithm for sparse regression and nonnegative least squares for numerical optimization. Analysis of 2,700 synthetic cancer genomes with and without noise demonstrates that SigProfilerAssignment outperforms four commonly used approaches for assigning mutational signatures. SigProfilerAssignment is freely available at https://github.com/AlexandrovLab/SigProfilerAssignment with a web implementation at https://cancer.sanger.ac.uk/signatures/assignment/.

Evidence-based suspicion and the prior probability of guilt in police interrogations.

OBJECTIVE: False confessions are prevalent in wrongful convictions, and much research has examined investigation factors and interrogation methods that can contribute to false confessions. However, not all these factors are under the control of the legal system, and improving the effectiveness of interrogation methods has a limited effect on evaluating the veracity of confessions. We suggest incorporating an important but often-neglected factor in interrogations: suspects' prior probability of guilt ("the prior," a Bayesian term meaning suspects' likelihood of being guilty before police conduct an interrogation). METHOD: By connecting interrogation practices to probability concepts, we discuss a gap in the literature between questions traditionally answered by lab research and a distinct question faced by the legal system. RESULTS: On the basis of our analysis, we argue that police should increase priors by collecting additional evidence to satisfy an evidence-based suspicion of guilt before interrogating suspects. CONCLUSIONS: Implementing the evidence-based suspicion practice can help police reduce false confessions, reallocate investigation time and resources, and assist prosecutors in building strong cases for trial. Likewise, researchers should expand the empirical and legal questions they ask and incorporate priors into their interrogation experiments to improve the generalizability of findings to the criminal justice system. (PsycInfo Database Record (c) 2023 APA, all rights reserved).

Uncovering novel mutational signatures by de novo extraction with SigProfilerExtractor.

Mutational signature analysis is commonly performed in cancer genomic studies. Here, we present SigProfilerExtractor, an automated tool for de novo extraction of mutational signatures, and benchmark it against another 13 bioinformatics tools by using 34 scenarios encompassing 2,500 simulated signatures found in 60,000 synthetic genomes and 20,000 synthetic exomes. For simulations with 5% noise, reflecting high-quality datasets, SigProfilerExtractor outperforms other approaches by elucidating between 20% and 50% more true-positive signatures while yielding 5-fold less false-positive signatures. Applying SigProfilerExtractor to 4,643 whole-genome- and 19,184 whole-exome-sequenced cancers reveals four novel signatures. Two of the signatures are confirmed in independent cohorts, and one of these signatures is associated with tobacco smoking. In summary, this report provides a reference tool for analysis of mutational signatures, a comprehensive benchmarking of bioinformatics tools for extracting signatures, and several novel mutational signatures, including one putatively attributed to direct tobacco smoking mutagenesis in bladder tissues.

The changing landscape of immune cells in the fetal mouse testis.

Fetal testis growth involves cell influx and extensive remodeling. Immediately after sex determination in mouse, macrophages enable normal cord formation and removal of inappropriately positioned cells. This study provides new information about macrophages and other immune cells after cord formation in fetal testes, including their density, distribution, and close cellular contacts. C57BL6J mouse testes from embryonic day (E) 13.5 to birth (post-natal day 0; PND0), were examined using immunofluorescence, immunohistochemistry, and RT-qPCR to identify macrophages (F4/80, CD206, MHCII), T cells (CD3), granulocytes/neutrophils (Ly6G), and germ cells (DDX4). F4/80+ cells were the most abundant, comprising 90% of CD45+ cells at E13.5 and declining to 65% at PND0. Changes in size, shape, and markers (CD206 and MHCII) documented during this interval align with the understanding that F4/80+ cells have different origins during embryonic life. CD3+ cells and F4/80-/MHCII+ were absent to rare until PND0. Ly6G+ cells were scarce at E13.5 but increased robustly by PND0 to represent half of the CD45+ cells. These immunofluorescence data were in accord with transcript analysis, which showed that immune marker mRNAs increased with testis age. F4/80+ and Ly6G+ cells were frequently inside cords adjacent to germ cells at E13.5 and E15.5. F4/80+ cells were often in clusters next to other immune cells. Macrophages inside cords at E13.5 and E15.5 (F4/80Hi/CD206+) were different from macrophages at PND0 (F4/80Dim/CD206-), indicating that they have distinct origins. This histological quantification coupled with transcript information identifies new cellular interactions for immune cells in fetal testis morphogenesis, and highlights new avenues for studies of their functional significance.

An international report on bacterial communities in esophageal squamous cell carcinoma.

The incidence of esophageal squamous cell carcinoma (ESCC) is disproportionately high in the eastern corridor of Africa and parts of Asia. Emerging research has identified a potential association between poor oral health and ESCC. One possible link between poor oral health and ESCC involves the alteration of the microbiome. We performed an integrated analysis of four independent sequencing efforts of ESCC tumors from patients from high- and low-incidence regions of the world. Using whole genome sequencing (WGS) and RNA sequencing (RNAseq) of ESCC tumors from 61 patients in Tanzania, we identified a community of bacteria, including members of the genera Fusobacterium, Selenomonas, Prevotella, Streptococcus, Porphyromonas, Veillonella and Campylobacter, present at high abundance in ESCC tumors. We then characterized the microbiome of 238 ESCC tumor specimens collected in two additional independent sequencing efforts consisting of patients from other high-ESCC incidence regions (Tanzania, Malawi, Kenya, Iran, China). This analysis revealed similar ESCC-associated bacterial communities in these cancers. Because these genera are traditionally considered members of the oral microbiota, we next explored whether there was a relationship between the synchronous saliva and tumor microbiomes of ESCC patients in Tanzania. Comparative analyses revealed that paired saliva and tumor microbiomes were significantly similar with a specific enrichment of Fusobacterium and Prevotella in the tumor microbiome. Together, these data indicate that cancer-associated oral bacteria are associated with ESCC tumors at the time of diagnosis and support a model in which oral bacteria are present in high abundance in both saliva and tumors of some ESCC patients.

The Impact of Activin A on Fetal Gonocytes: Chronic Versus Acute Exposure Outcomes.

Activin A, a TGFß superfamily member, is important for normal testis development through its actions on Sertoli cell development. Our analyses of altered activin A mouse models indicated gonocyte abnormalities, implicating activin A as a key determinant of early germline formation. Whether it acts directly or indirectly on germ cells is not understood. In humans, the fetal testis may be exposed to abnormally elevated activin A levels during preeclampsia, maternal infections, or following ingestion of certain medications. We hypothesized that this may impact fetal testis development and ultimately affect adult fertility. Germ cells from two mouse models of altered activin bioactivity were analysed. RNA-Seq of gonocytes purified from E13.5 and E15.5 Inhba KO mice (activin A subunit knockout) identified 46 and 44 differentially expressed genes (DEGs) respectively, and 45 in the E13.5 Inha KO (inhibin alpha subunit knockout; increased activin A) gonocytes. To discern direct effects of altered activin bioactivity on germline transcripts, isolated E13.5 gonocytes were cultured for 24h with activin A or with the activin/Nodal/TGFß inhibitor, SB431542. Gonocytes responded directly to altered signalling, with activin A promoting a more differentiated transcript profile (increased differentiation markers Dnmt3l, Nanos2 and Piwil4; decreased early germ cell markers Kit and Tdgf1), while SB431542 had a reciprocal effect (decreased Nanos2 and Piwil4; increased Kit). To delineate direct and indirect effects of activin A exposure on gonocytes, whole testes were cultured 48h with activin A or SB431542 and collected for histological and transcript analyses, or EdU added at the end of culture to measure germ and Sertoli cell proliferation using flow cytometry. Activin increased, and SB431542 decreased, Sertoli cell proliferation. SB431542-exposure resulted in germ cells escaping mitotic arrest. Analysis of FACS-isolated gonocytes following whole testis culture showed SB431542 increased the early germ cell marker Kit, however there was a general reduction in the impact of altered activin A bioavailability in the normal somatic cell environment. This multifaceted approach identifies a capacity for activin A to directly influence fetal germ cell development, highlighting the potential for altered activin A levels in utero to increase the risk of testicular pathologies that arise from impaired germline maturation.

Mutational signatures in esophageal squamous cell carcinoma from eight countries with varying incidence.

Esophageal squamous cell carcinoma (ESCC) shows remarkable variation in incidence that is not fully explained by known lifestyle and environmental risk factors. It has been speculated that an unknown exogenous exposure(s) could be responsible. Here we combine the fields of mutational signature analysis with cancer epidemiology to study 552 ESCC genomes from eight countries with varying incidence rates. Mutational profiles were similar across all countries studied. Associations between specific mutational signatures and ESCC risk factors were identified for tobacco, alcohol, opium and germline variants, with modest impacts on mutation burden. We find no evidence of a mutational signature indicative of an exogenous exposure capable of explaining differences in ESCC incidence. Apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like (APOBEC)-associated mutational signatures single-base substitution (SBS)2 and SBS13 were present in 88% and 91% of cases, respectively, and accounted for 25% of the mutation burden on average, indicating that APOBEC activation is a crucial step in ESCC tumor development.

Loss of the long non-coding RNA OIP5-AS1 exacerbates heart failure in a sex-specific manner.

Long non-coding RNAs (lncRNAs) have been demonstrated to influence numerous biological processes, being strongly implicated in the maintenance and physiological function of various tissues including the heart. The lncRNA OIP5-AS1 (1700020I14Rik/Cyrano) has been studied in several settings; however its role in cardiac pathologies remains mostly uncharacterized. Using a series of in vitro and ex vivo methods, we demonstrate that OIP5-AS1 is regulated during cardiac development in rodent and human models and in disease settings in mice. Using CRISPR, we engineered a global OIP5-AS1 knockout (KO) mouse and demonstrated that female KO mice develop exacerbated heart failure following cardiac pressure overload (transverse aortic constriction [TAC]) but male mice do not. RNA-sequencing of wild-type and KO hearts suggest that OIP5-AS1 regulates pathways that impact mitochondrial function. Thus, these findings highlight OIP5-AS1 as a gene of interest in sex-specific differences in mitochondrial function and development of heart failure.

Thyroid MALT lymphoma: self-harm to gain potential T-cell help.

The development of extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT) is driven by chronic inflammatory responses and acquired genetic changes. To investigate its genetic bases, we performed targeted sequencing of 93 genes in 131 MALT lymphomas including 76 from the thyroid. We found frequent deleterious mutations of TET2 (86%), CD274 (53%), TNFRSF14 (53%), and TNFAIP3 (30%) in thyroid MALT lymphoma. CD274 was also frequently deleted, together with mutation seen in 68% of cases. There was a significant association between CD274 mutation/deletion and TNFRSF14 mutation (p = 0.001). CD274 (PD-L1) and TNFRSF14 are ligands for the co-inhibitory receptor PD1 and BTLA on T-helper cells, respectively, their inactivation may free T-cell activities, promoting their help to malignant B-cells. In support of this, both the proportion of activated T-cells (CD4+CD69+/CD4+) within the proximity of malignant B-cells, and the level of transformed blasts were significantly higher in cases with CD274/TNFRSF14 genetic abnormalities than those without these changes. Both CD274 and TNFRSF14 genetic changes were significantly associated with Hashimoto's thyroiditis (p = 0.01, p = 0.04, respectively), and CD274 mutation/deletion additionally associated with increased erythrocyte sedimentation rate (p = 0.0001). In conclusion, CD274/TNFRSF14 inactivation in thyroid MALT lymphoma B-cells may deregulate their interaction with T-cells, promoting co-stimulations and impairing peripheral tolerance.

Cannabis, Cannabidiol Oils and Tetrahydrocannabinol-What Do Veterinarians Need to Know?

As cannabis-derived products have become more available, veterinarians are seeing more cases of toxicosis. In addition, animal owners are having an increasing interest in using these products for their pets. This review looks at the situation in Europe and North America, the different types of cannabis and cannabis-derived products with historical examples of use in animals, and the cannabis industry. The existing regulatory framework for use in humans and animals as medicines and/or supplements was examined. Finally, a review of the clinical indications for which medicinal cannabis is authorised, a discussion of toxicosis, and recommendations and warnings around medical cannabis use are presented.

Deletion of Trim28 in committed adipocytes promotes obesity but preserves glucose tolerance.

The effective storage of lipids in white adipose tissue (WAT) critically impacts whole body energy homeostasis. Many genes have been implicated in WAT lipid metabolism, including tripartite motif containing 28 (Trim28), a gene proposed to primarily influence adiposity via epigenetic mechanisms in embryonic development. However, in the current study we demonstrate that mice with deletion of Trim28 specifically in committed adipocytes, also develop obesity similar to global Trim28 deletion models, highlighting a post-developmental role for Trim28. These effects were exacerbated in female mice, contributing to the growing notion that Trim28 is a sex-specific regulator of obesity. Mechanistically, this phenotype involves alterations in lipolysis and triglyceride metabolism, explained in part by loss of Klf14 expression, a gene previously demonstrated to modulate adipocyte size and body composition in a sex-specific manner. Thus, these findings provide evidence that Trim28 is a bona fide, sex specific regulator of post-developmental adiposity and WAT function.

Does Perpetrator Occupation Affect Classification of Sexual Assault?

Classification of sexual assault varies based on the characteristics of the victim, perpetrator, and event. However, most studies focus on the individual characteristics of participants asked to classify the assault, the victims' characteristics, or the event; few have examined variations in the perpetrator. Therefore, this study examined whether the occupation of the perpetrator affected the classification of the event as sexual assault. Participants included a primarily White female sample of undergraduate students (n = 401) at a south-Atlantic university. They completed a paper-and-pencil survey containing an ambiguous sexual assault encounter where the occupation of the perpetrator (athlete, reporter, college student, or politician) was randomly varied. Participants were asked to classify whether the encounter was sexual assault and what an appropriate punishment might be. Additional measures included the Illinois Rape Myth Acceptance Scale (IRMA) and questions about personal experience with sexual assault. Overall, most participants labeled the encounter as sexual assault, indicating that college students are aware of the legal parameters for providing consent. However, this classification occurred differentially based on the occupation of the perpetrator. Participants were most likely to label the athlete as committing sexual assault, followed by the college student, politician, and reporter, respectively. Females and those who reject rape myths were more likely to label the scenario as sexual assault. A "personal apology" and a "sexual rehabilitation program" were the most common punishments selected for the perpetrator. These findings highlight potential concerns and the need for additional training when college students adjudicate sexual assault reports and determine appropriate consequences.

Mutagenicity of 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N-OH-PhIP) in human TP53 knock-in (Hupki) mouse embryo fibroblasts.

2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a possible human carcinogen formed in cooked fish and meat. PhIP is bioactivated by cytochrome P450 enzymes to form 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N-OH-PhIP), a genotoxic metabolite that reacts with DNA leading to the mutation-prone DNA adduct N-(deoxyguanosin-8-yl)-PhIP (dG-C8-PhIP). Here, we studied N-OH-PhIP-induced whole genome mutagenesis in human TP53 knock-in (Hupki) mouse embryo fibroblasts (HUFs) immortalised and subjected to whole genome sequencing (WGS). In addition, mutagenicity of N-OH-PhIP in TP53 and the lacZ reporter gene were assessed. TP53 mutant frequency in HUF cultures treated with N-OH-PhIP (2.5 µM for 24 h, n = 90) was 10% while no TP53 mutations were found in untreated controls (DMSO for 24 h, n = 6). All N-OH-PhIP-induced TP53 mutations occurred at G:C base pairs with G > T/C > A transversions accounting for 58% of them. TP53 mutations characteristic of those induced by N-OH-PhIP have been found in human tumours including breast and colorectal, which are cancer types that have been associated with PhIP exposure. LacZ mutant frequency increased 25-fold at 5 µM N-OH-PHIP and up to ~350 dG-C8-PhIP adducts/108 nucleosides were detected by ultra-performance liquid chromatography-electrospray ionisation multistage scan mass spectrometry (UPLC-ESI-MS3) at this concentration. In addition, a WGS mutational signature defined by G > T/C > A transversions was present in N-OH-PhIP-treated immortalised clones, which showed similarity to COSMIC SBS4, 18 and 29 signatures found in human tumours.

Mutagenicity of acrylamide and glycidamide in human TP53 knock-in (Hupki) mouse embryo fibroblasts.

Acrylamide is a suspected human carcinogen formed during high-temperature cooking of starch-rich foods. It is metabolised by cytochrome P450 2E1 to its reactive metabolite glycidamide, which forms pre-mutagenic DNA adducts. Using the human TP53 knock-in (Hupki) mouse embryo fibroblasts (HUFs) immortalisation assay (HIMA), acrylamide- and glycidamide-induced mutagenesis was studied in the tumour suppressor gene TP53. Selected immortalised HUF clones were also subjected to next-generation sequencing to determine mutations across the whole genome. The TP53-mutant frequency after glycidamide exposure (1.1 mM for 24 h, n = 198) was 9% compared with 0% in cultures treated with acrylamide [1.5 (n = 24) or 3 mM (n = 6) for 48 h] and untreated vehicle (water) controls (n = 36). Most glycidamide-induced mutations occurred at adenines with A > T/T > A and A > G/T > C mutations being the most common types. Mutations induced by glycidamide occurred at specific TP53 codons that have also been found to be mutated in human tumours (i.e., breast, ovary, colorectal, and lung) previously associated with acrylamide exposure. The spectrum of TP53 mutations was further reflected by the mutations detected by whole-genome sequencing (WGS) and a distinct WGS mutational signature was found in HUF clones treated with glycidamide that was again characterised by A > G/T > C and A > T/T > A mutations. The WGS mutational signature showed similarities with COSMIC mutational signatures SBS3 and 25 previously found in human tumours (e.g., breast and ovary), while the adenine component was similar to COSMIC SBS4 found mostly in smokers' lung cancer. In contrast, in acrylamide-treated HUF clones, only culture-related background WGS mutational signatures were observed. In summary, the results of the present study suggest that glycidamide may be involved in the development of breast, ovarian, and lung cancer.

Disparate Effects of Diabetes and Hyperlipidemia on Experimental Kidney Disease.

It is well established that diabetes is the major cause of chronic kidney disease worldwide. Both hyperglycemia, and more recently, advanced glycation endproducts, have been shown to play critical roles in the development of kidney disease. Moreover, the renin-angiotensin system along with growth factors and cytokines have also been shown to contribute to the onset and progression of diabetic kidney disease; however, the role of lipids in this context is poorly characterized. The current study aimed to compare the effect of 20 weeks of streptozotocin-induced diabetes or western diet feeding on kidney disease in two different mouse strains, C57BL/6 mice and hyperlipidemic apolipoprotein (apo) E knockout (KO) mice. Mice were fed a chow diet (control), a western diet (21% fat, 0.15% cholesterol) or were induced with streptozotocin-diabetes (55 mg/kg/day for 5 days) then fed a chow diet and followed for 20 weeks. The induction of diabetes was associated with a 3-fold elevation in glycated hemoglobin and an increase in kidney to body weight ratio regardless of strain (p < 0.0001). ApoE deficiency significantly increased plasma cholesterol and triglyceride levels and feeding of a western diet exacerbated these effects. Despite this, urinary albumin excretion (UAE) was elevated in diabetic mice to a similar extent in both strains (p < 0.0001) but no effect was seen with a western diet in either strain. Diabetes was also associated with extracellular matrix accumulation in both strains, and western diet feeding to a lesser extent in apoE KO mice. Consistent with this, an increase in renal mRNA expression of the fibrotic marker, fibronectin, was observed in diabetic C57BL/6 mice (p < 0.0001). In summary, these studies demonstrate disparate effects of diabetes and hyperlipidemia on kidney injury, with features of the diabetic milieu other than lipids suggested to play a more prominent role in driving renal pathology.

Evidence that activin A directly modulates early human male germline differentiation status.

Disrupted fetal germline development underpins testicular germ cell neoplasia, which is increasing worldwide. The complex signaling milieu during normal testis development includes TGFß superfamily ligands; this study tests the hypothesis that, activin A, a TGFß superfamily member, can influence gonocyte development. The human seminoma-derived cell line, TCam-2, a model of fetal gonocytes, was cultured with activin A (1.25-25 ng/mL) for 48 h, or with 5 ng/mL activin A for short- (6, 24, and 48 h) and long-term (13 days) exposures, and downstream targets measured by qRT-PCR. Transcripts that exhibited significant dose-dependent responses to activin A included the early germ cell markers KIT, NODAL, and CRIPTO (NODALl co-receptor and activin inhibitor) which all increased and the differentiation marker DNMT3L which decreased. After 48 h, KIT, NODAL, and CRIPTO levels were significantly higher, while the differentiation marker NANOS2 was significantly lower. Interestingly, activin A exposure also significantly reduced both transcript and protein levels of the PIWI/piRNA pathway component DNMT3L. Because TCam-2 cells produce the activin inhibitor CRIPTO, CRIPTO was reduced using siRNA prior to activin A exposure. This selectively increased KIT in response to activin A. Other ligands present in the fetal testis (BMP4, FGF9, TGFß1, and TGFß2) induced distinct effects on germline marker expression. This study showed that activin A can directly modulate germline markers in this human gonocyte-like cell, promoting a less-differentiated phenotype. Additional findings indicate evidence of signaling crosstalk between activin A and NODAL, leading to target-specific effects on gonocyte differentiation.