Mycoplasma DnaK expression increases cancer development in vivo upon DNA damage.

Well-controlled repair mechanisms are involved in the maintenance of genomic stability, and their failure can precipitate DNA abnormalities and elevate tumor risk. In addition, the tumor microenvironment, enriched with factors inducing oxidative stress and affecting cell cycle checkpoints, intensifies DNA damage when repair pathways falter. Recent research has unveiled associations between certain bacteria, including Mycoplasmas, and various cancers, and the causative mechanism(s) are under active investigation. We previously showed that Mycoplasma fermentans DnaK, an HSP70 family chaperone protein, hampers the activity of proteins like PARP1 and p53, crucial for genomic integrity. Moreover, our analysis of its interactome in human cancer cell lines revealed DnaK's engagement with several components of DNA-repair machinery. Finally, in vivo experiments performed in our laboratory using a DnaK knock-in mouse model generated by our group demonstrated that DnaK exposure led to increased DNA copy number variants, indicative of genomic instability. We present here evidence that expression of DnaK is linked to increased i) incidence of tumors in vivo upon exposure to urethane, a DNA damaging agent; ii) spontaneous DNA damage ex vivo; and iii) expression of proinflammatory cytokines ex vivo, variations in reactive oxygen species levels, and increased ß-galactosidase activity across tissues. Moreover, DnaK was associated with increased centromeric instability. Overall, these findings highlight the significance of Mycoplasma DnaK in the etiology of cancer and other genetic disorders providing a promising target for prevention, diagnostics, and therapeutics.

Inflammatory Myositis Following Statin Use in a Patient With Untreated Hypothyroidism.

Inflammatory myositis (IM) presents a diagnostic challenge due to its multifaceted etiology and varying clinical presentations. This case involves a 55-year-old male with asymptomatic hypothyroidism, recent statin use, and rapidly progressing proximal muscle weakness. He presented with profound weakness in the upper and lower extremities, severely impairing his daily activities. The patient's medical history included recent hospitalizations for idiopathic interstitial lung disease, myopericarditis, and pneumonia, adding complexity to his condition. Laboratory findings revealed elevated serum muscle enzymes, notably creatine kinase, indicating muscle damage and rhabdomyolysis. Serological testing confirmed the absence of myositis-specific antibodies and anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) antibodies. The patient was eventually diagnosed with IM and rhabdomyolysis, likely secondary to statin use or hypothyroidism. Treatment with methylprednisolone, levothyroxine, and discontinuation of atorvastatin led to rapid improvements in AST levels and overall muscle strength. This case highlights the challenges of managing IM and emphasizes the importance of assessing thyroid function before initiating lipid-lowering therapy.

Correlation of HIV-Induced Neuroinflammation and Synaptopathy with Impairment of Learning and Memory in Mice with HAND.

Over 38 million people worldwide are living with HIV/AIDS, and more than half of them are affected by HIV-associated neurocognitive disorders (HAND). Such disorders are characterized by chronic neuroinflammation, neurotoxicity, and central nervous system deterioration, which lead to short- or long-term memory loss, cognitive impairment, and motor skill deficits that may show gender disparities. However, the underlying mechanisms remain unclear. Our previous study suggested that HIV-1 infection and viral protein R (Vpr) upregulate the SUR1-TRPM4 channel associated with neuroinflammation, which may contribute to HAND. The present study aimed to explore this relationship in a mouse model of HAND. This study employed the HIV transgenic Tg26 mouse model, comparing Tg26 mice with wildtype mice in various cognitive behavioral and memory tests, including locomotor activity tests, recognition memory tests, and spatial learning and memory tests. The study found that Tg26 mice exhibited impaired cognitive skills and reduced learning abilities compared to wildtype mice, particularly in spatial memory. Interestingly, male Tg26 mice displayed significant differences in spatial memory losses (p < 0.001), while no significant differences were identified in female mice. Consistent with our early results, SUR1-TRPM4 channels were upregulated in Tg26 mice along with glial fibrillary acidic protein (GFAP) and aquaporin 4 (AQP4), consistent with reactive astrocytosis and neuroinflammation. Corresponding reductions in neurosynaptic responses, as indicated by downregulation of Synapsin-1 (SYN1) and Synaptophysin (SYP), suggested synaptopathy as a possible mechanism underlying cognitive and motor skill deficits. In conclusion, our study suggests a possible relationship between SUR1-TRPM4-mediated neuroinflammation and synaptopathy with impairments of learning and memory in mice with HAND. These findings could help to develop new therapeutic strategies for individuals living with HAND.

Mycoplasma DnaK increases DNA copy number variants in vivo.

The human microbiota affects critical cellular functions, although the responsible mechanism(s) is still poorly understood. In this regard, we previously showed that Mycoplasma fermentans DnaK, an HSP70 chaperone protein, hampers the activity of important cellular proteins responsible for DNA integrity. Here, we describe a novel DnaK knock-in mouse model generated in our laboratory to study the effect of M. fermentans DnaK expression in vivo. By using an array-based comparative genomic hybridization assay, we demonstrate that exposure to DnaK was associated with a higher number of DNA copy number variants (CNVs) indicative of unbalanced chromosomal alterations, together with reduced fertility and a high rate of fetal abnormalities. Consistent with their implication in genetic disorders, one of these CNVs caused a homozygous Grid2 deletion, resulting in an aberrant ataxic phenotype that recapitulates the extensive biallelic deletion in the Grid2 gene classified in humans as autosomal recessive spinocerebellar ataxia 18. Our data highlight a connection between components of the human urogenital tract microbiota, namely Mycoplasmas, and genetic abnormalities in the form of DNA CNVs, with obvious relevant medical, diagnostic, and therapeutic implications.

A Comparison of Lymphoid and Myeloid Cells Derived from Human Hematopoietic Stem Cells Xenografted into NOD-Derived Mouse Strains.

Humanized mice are an invaluable tool for investigating human diseases such as cancer, infectious diseases, and graft-versus-host disease (GvHD). However, it is crucial to understand the strengths and limitations of humanized mice and select the most appropriate model. In this study, we describe the development of the human lymphoid and myeloid lineages using a flow cytometric analysis in four humanized mouse models derived from NOD mice xenotransplanted with CD34+ fetal cord blood from a single donor. Our results showed that all murine strains sustained human immune cells within a proinflammatory environment induced by GvHD. However, the Hu-SGM3 model consistently generated higher numbers of human T cells, monocytes, dendritic cells, mast cells, and megakaryocytes, and a low number of circulating platelets showing an activated profile when compared with the other murine strains. The hu-NOG-EXL model had a similar cell development profile but a higher number of circulating platelets with an inactivated state, and the hu-NSG and hu-NCG developed low frequencies of immune cells compared with the other models. Interestingly, only the hu-SGM3 and hu-EXL models developed mast cells. In conclusion, our findings highlight the importance of selecting the appropriate humanized mouse model for specific research questions, considering the strengths and limitations of each model and the immune cell populations of interest.

Humanized Mice for the Study of Dengue Disease Pathogenesis: Biological Assays.

Dengue is one of the most prevalent infectious diseases around the world, present in all continents and mainly affecting developing countries. With few tools to fight and study this disease, it is imperative to have reliable animal models that not only recapitulate human disease but also contain human components to understand the pathogenic mechanism and immune responses, allowing the development of new treatments and vaccines against dengue. Humanized mice are a significant advance in the development of in vivo models to understanding the relation of the human immune system and target organs such as the liver during the infection by dengue virus, allowing basic and preclinical research. In this chapter, we describe the use of humanized NSG mice (huNSG) for the study of dengue disease. The first model describes reconstitution of the human immune system by transplanting human CD34+ stem cells in newborn or adult NSG mice. The second model combines the reconstitution with CD34+ stem cells with the transplant of human primary hepatocytes. This dual reconstituted animal will have two of the major players involved in the development of dengue infection. However, there are still more biological components missing in this model for dengue, but researchers continue working to improve the huNSG model to reconstitute other human components.

Cigarette smoke and nicotine effects on behavior in HIV transgenic rats.

HIV-related neurocognitive impairment can be worsened by cigarette smoking and be more severe in women. Therefore, we analyzed the effects of sex on behavioral function in HIV transgenic (Tg) rats that were exposed to either nicotine alone, to smoke from either nicotine-containing or nicotine-free cigarettes, or non-exposed. The animals were then assessed on the open field test for the total distance traveled and for the fraction of the total distance traveled and the total time spent in the center of the field, and the results then compared to WT rats subjected to the same exposures and testing. Higher total distances indicate greater locomotor activity and a higher center field measures imply a lower anxiety state. Total distances were overall higher for female and for Tg rats exposed to nicotine-free CS. Also, the total distance and both center field measures were overall higher for female rats in the control and nicotine-free CS-exposed groups. This was observed specifically for WT females as compared to WT males and, for the center field measures, for WT females as compared to Tg males. No genotype or sex-related differences were found for rats in the nicotine-free cigarette smoke (CS) and nicotine-containing CS exposed groups. Therefore, nicotine exposure did not impact genotype- and sex-related differences in motor responses and anxiety levels that were found in the control state. However, exposure to the non-nicotine components of CS resulted in locomotor activation in the presence of the HIV genes and was anxiogenic in WT and Tg male animals.

Lipid lowering in patients 75 years and older.

More than twenty years ago, knowledge about the importance of cholesterol absorption and the potential therapeutic effect of its inhibition led to the discovery and clinical application of the first and only cholesterol absorption inhibitor to date - ezetimibe. Since then, ezetimibe has become a well-recognized player in lipid-lowering therapy. Recent findings of IMPROVE-IT and EWTOPIA 75 imply that elderly patients over the age of 75 years in particular benefit from ezetimibe. This review summarizes the evidence, discusses the possible underlying pathophysiological mechanisms and calls for a change in future dyslipidemia guidelines.

APOL1 Genetic Variants Are Associated With Increased Risk of Coronary Atherosclerotic Plaque Rupture in the Black Population.

[Figure: see text].

Everyday experiences of post-diagnosis life with dementia: A co-produced photography study.

There has been surprisingly little research capturing people's everyday lives in the early years following a diagnosis of dementia. This project was co-produced by three people with dementia and two university researchers. The co-researchers with dementia formulated the aims of this project as: (1) to explore post-diagnosis life with dementia and (2) to use data collection methods as a form of peer support and confidence building for the participants. The intent was to provide the opportunity to learn new skills and support participants to share their experiences without putting them on the spot. Five participants with recent diagnoses received a photography lesson and cameras to take photographs of their everyday lives. This was followed with a focus group in which the photographs were discussed. The participants used their photographs to explain: (1) the differences between their past and present with dementia, (2) how dementia affected their thought processes, (3) pets and grandchildren as facilitators of reciprocal joy and support, (4) the emotional effects of living with a dementia diagnosis, and (5) the solace and stability of nature in a changing world. The participants' creative use of photography supported them to express the complex emotions felt after a diagnosis of dementia and they reported the benefit of doing this in an environment with peers going through the same experiences. The role of the co-researchers with dementia was the key to the success of this project, drawing on their own experiences to design the project and support the participants. Future research should draw on the experiences of people with dementia to design research projects and care interventions.

The HIV-1 Transgenic Rat: Relevance for HIV Noninfectious Comorbidity Research.

HIV noninfectious comorbidities (NICMs) are a current healthcare challenge. The situation is further complicated as there are very few effective models that can be used for NICM research. Previous research has supported the use of the HIV-1 transgenic rat (HIV-1TGR) as a model for the study of HIV/AIDS. However, additional studies are needed to confirm whether this model has features that would support NICM research. A demonstration of the utility of the HIV-1TGR model would be to show that the HIV-1TGR has cellular receptors able to bind HIV proteins, as this would be relevant for the study of cell-specific tissue pathology. In fact, an increased frequency of HIV receptors on a specific cell type may increase tissue vulnerability since binding to HIV proteins would eventually result in cell dysfunction and death. Evidence suggests that observations of selective cellular vulnerability in this model are consistent with some specific tissue vulnerabilities seen in NICMs. We identified CXCR4-expressing cells in the brain, while specific markers for neuronal degeneration demonstrated that the same neural types were dying. We also confirm the presence of gp120 and Tat by immunocytochemistry in the spleen, as previously reported. However, we observed very rare positive cells in the brain. This underscores the point that gp120, which has been reported as detected in the sera and CSF, is a likely source to which these CXCR4-positive cells are exposed. This alternative appears more probable than the local production of gp120. Further studies may indicate some level of local production, but that will not eliminate the role of receptor-mediated pathology. The binding of gp120 to the CXCR4 receptor on neurons and other neural cell types in the HIV-1TGR can thus explain the phenomena of selective cell death. Selective cellular vulnerability may be a contributing factor to the development of NICMs. Our data indicate that the HIV-1TGR can be an effective model for the studies of HIV NICMs because of the difference in the regional expression of CXCR4 in rat tissues, thus leading to specific organ pathology. This also suggests that the model can be used in the development of therapeutic options.

Characterization of CXCR5+ CD8+ T-cells in humanized NSG mice.

Humanized NOD/SCID/IL-2 receptor γ-chainnull (huNSG) mice recapitulate some features of human T-cell populations that can be exploited in basic and pre-clinical research. CXCR5+ T CD8+ T-cells play an important role in the control of viral infections and tumors. Indeed, they have been associated with low-level HIV replication, making them a possible novel correlate of protection, and potentially useful in the eradication of HIV reservoirs. Here, by flow cytometry, we evaluated the reconstitution of CXCR5+ CD8+ T-cells in huNSG mice engrafted with CD34+ hematopoietic stem cells. This population was readily generated in huNSG mice, and where particularly confined to spleen and lymph nodes. These cells exhibited a follicular-like phenotype, with expression of Programmed Death (PD)-1, Inducible T-cell costimulatory (ICOS), and absence of CCR7. Moreover, CXCR5+ CD8+ T-cells had a higher expression of interleukin (IL)-21 and a higher cytotoxic potential compared with CXCR5- cells. HIV infection did not affect the frequencies of CXCR5+ CD8+ T-cells in secondary lymphoid organs. Finally, taking advantage of the high proportion of naïve T-cells in huNSG mice, we evaluated the in vitro response of splenic T-cells to the follicular profile-polarizing cytokines Transforming Growth Factor (TGF)-ß1 and IL-23. After in vitro treatment, there was an increase in CXCR5+ CD8+ T-cells, which exhibited high levels of PD-1, CD40 L and low expression of CCR7. Thus, there is a reconstitution of CXCR5+ CD8+ T-cells in huNSG mice, supporting the use of this model for exploring the biology and role of this cell population in healthy and diseased conditions.

Chronic, Acute, and Reactivated HIV Infection in Humanized Immunodeficient Mouse Models.

Humanized NOD/SCID/IL-2 receptor γ-chainnull mice recapitulate some features of human immunity, which can be exploited in basic and pre-clinical research on infectious diseases. Described here are three models of humanized immunodeficient mice for studying the dynamics of HIV infection. The first is based on the intrahepatic injection of CD34+ hematopoietic stem cells in newborn mice, which allows for the reconstitution of several blood and lymphoid tissue-confined cells, followed by infection with a reference HIV strain. This model allows monitoring for up to 36 weeks post-infection and is hence called the chronic model. The second and third models are referred to as the acute and reactivation models, in which peripheral blood mononuclear cells are intraperitoneally injected in adult mice. In the acute model, cells from a healthy donor are engrafted through the intraperitoneal route, followed by infection with a reference HIV strain. Finally, in the reactivation model, cells from an HIV-infected donor under antiretroviral therapy are engrafted via the intraperitoneal route. In this case, a drug-free environment in the mouse allows for virus reactivation and an increase in viral load. The protocols provided here describe the conventional experimental approach for humanized, immunodeficient mouse models of HIV infection.

HIV Replication in Humanized IL-3/GM-CSF-Transgenic NOG Mice.

The development of mouse models that mimic the kinetics of Human Immunodeficiency Virus (HIV) infection is critical for the understanding of the pathogenesis of disease and for the design of novel therapeutic strategies. Here, we describe the dynamics of HIV infection in humanized NOD/Shi-scid-IL2rγnull (NOG) mice bearing the human genes for interleukin (IL)-3 and granulocyte-macrophage colony-stimulating factor (GM-CSF) (NOG-EXL mice). The kinetics of viral load, as well as the frequencies of T-cells, B-cells, Natural killer cells (NK), monocytes, and dendritic cells in blood and secondary lymphoid organs were evaluated throughout the time of infection. In comparison with a non-transgenic humanized mouse (NSG) strain, lymphoid and myeloid populations were more efficiently engrafted in humanized NOG-EXL mice, both in peripheral blood and lymphoid tissues. In addition, HIV actively replicated in humanized NOG-EXL mice, and infection induced a decrease in the percentage of CD4⁺ T-cells, inversion of the CD4:CD8 ratio, and changes in some cell populations, such as monocytes and dendritic cells, that recapitulated those found in human natural infection. Thus, the humanized IL-3/GM-CSF-transgenic NOG mouse model is suitable for the study of the dynamics of HIV infection and provides a tool for basic and preclinical studies.

HIVAN associated tubular pathology with reference to ER stress, mitochondrial changes, and autophagy.

Human immunodeficiency virus associated nephropathy (HIVAN) is a unique form of a renal parenchymal disorder. This disease and its characteristics can be accredited to incorporation of DNA and mRNA of human immunodeficiency virus type 1 into the renal parenchymal cells. A proper understanding of the intricacies of HIVAN and the underlying mechanisms associated with renal function and disorders is vital for the potential development of a reliable treatment for HIVAN. Specifically, the renal tubule segment of the kidney is characterized by its transport capabilities and its ability to reabsorb water and salts into the blood. However, the segment is also known for certain disorders, such as renal tubular epithelial cell infection and microcyst formation, which are also closely linked to HIVAN. Furthermore, certain organelles, like the endoplasmic reticulum (ER), mitochondria, and lysosome, are vital for certain underlying mechanisms in kidney cells. A paradigm of the importance of said organelles can be seen in documented cases of HIVAN where the renal disorder results increased ER stress due to HIV viral propagation. This balance can be restored through the synthesis of secretory proteins, but, in return, the secretion requires more energy; therefore, there is a noticeable increase in mitochondrial stress. The increased ER changes and mitochondrial stress will greatly upregulate the process of autophagy, which involves the cell's lysosomes. In conjunction, we found that ER stress and mitochondrial changes are associated in the Tg26 animal model of HIVAN. The aim of our review is to consolidate current knowledge of important mechanisms in HIVAN, specifically related to the renal tubules' association with ER stress, mitochondrial changes and autophagy. Although the specific regulatory mechanism detailing the cross-talk between the various organelles is unknown in HIVAN, the continued research in this field may potentially shed light on a possible improved treatment for HIVAN.

A20 deficiency in multipotent progenitors perturbs quiescence of hematopoietic stem cells.

Inflammatory signals have been shown to play a critical role in controlling the maintenance and functions of hematopoietic stem cells (HSCs). While the significance of inflammation in hematopoiesis has begun to unfold, molecular mechanisms and players that govern this mode of HSC regulation remain largely unknown. The E3 ubiquitin ligase A20 has been considered as a central gatekeeper of inflammation. Here, we have specifically depleted A20 in multi-potent progenitors (MPPs) and studied its impact on hematopoiesis. Our data suggest that lack of A20 in Flt3+ progenitors causes modest alterations in hematopoietic differentiation. Analysis of hematopoietic stem and progenitor cell (HSPC) pool revealed alterations in HSPC subsets including, HSCs, MPP1, MPP2, MPP3 and MPP4. Interestingly, A20 deficiency in MPPs caused loss of HSC quiescence and compromised long-term hematopoietic reconstitution. Mechanistic studies identified that A20 deficiency caused elevated levels of Interferon-γ signaling and downregulation of p57 in HSCs. In essence, these studies identified A20 as a key regulator of HSC quiescence and cell fate decisions.

Progress and perspectives in plant sterol and plant stanol research.

Current evidence indicates that foods with added plant sterols or stanols can lower serum levels of low-density lipoprotein cholesterol. This review summarizes the recent findings and deliberations of 31 experts in the field who participated in a scientific meeting in Winnipeg, Canada, on the health effects of plant sterols and stanols. Participants discussed issues including, but not limited to, the health benefits of plant sterols and stanols beyond cholesterol lowering, the role of plant sterols and stanols as adjuncts to diet and drugs, and the challenges involved in measuring plant sterols and stanols in biological samples. Variations in interindividual responses to plant sterols and stanols, as well as the personalization of lipid-lowering therapies, were addressed. Finally, the clinical aspects and treatment of sitosterolemia were reviewed. Although plant sterols and stanols continue to offer an efficacious and convenient dietary approach to cholesterol management, long-term clinical trials investigating the endpoints of cardiovascular disease are still lacking.

Effect of bioactive glass-containing resin composite on dentin remineralization.

OBJECTIVES: The purpose of this study was to evaluate the effect of bioactive glass (BAG)-containing composite on dentin remineralization. METHODS: Sixty-six dentin disks with 3 mm thickness were prepared from thirty-three bovine incisors. The following six experimental groups were prepared according to type of composite (control and experimental) and storage solutions (simulated body fluid [SBF] and phosphate-buffered saline [PBS]): 1 (undemineralized); 2 (demineralized); 3 (demineralized with control composite in SBF); 4 (demineralized with control composite in PBS); 5 (demineralized with experimental composite in SBF); and 6 (demineralized with experimental composite in PBS). BAG65S (65% Si, 31% Ca, and 4% P) was prepared via the sol-gel method. The control composite was made with a 50:50 Bis-GMA:TEGDMA resin matrix, 57 wt% strontium glass, and 15 wt% aerosol silica. The experimental composite had the same resin and filler, but with 15 wt% BAG65S replacing the aerosol silica. For groups 3-6, composite disks (20 × 10 × 2 mm) were prepared and approximated to the dentin disks and stored in PBS or SBF for 2 weeks. Micro-hardness test, attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR) and field-emission scanning electron microscopy (FE-SEM) was investigated. RESULTS: The BAG-containing composite significantly increased the micro-hardness of the adjacent demineralized dentin. ATR-FTIR revealed calcium phosphate peaks on the surface of the groups which used BAG-containing composite. FE-SEM revealed surface deposits partially occluding the dentin surface. No significant difference was found between SBF and PBS storage. CLINICAL SIGNIFICANCE: Bioactive glass is a potentially useful material for remineralization of tooth structure. When incorporated into a resin composite, it may aid in remineralizing the adjacent demineralized dentin, thus preventing further destruction of the tooth. CONCLUSION: BAG-containing composites placed in close proximity can partially remineralize adjacent demineralized dentin.