BACKGROUND: Toxoplasma gondii is an intracellular protozoan parasite that is widely distributed in humans and warm-blooded animals. T. gondii chronic infections can cause toxoplasmic encephalopathy, adverse pregnancy, and male reproductive disorders. In male reproduction, the main function of the testis is to provide a stable place for spermatogenesis and immunological protection. The disorders affecting testis tissue encompass abnormalities in the germ cell cycle, spermatogenic retardation, or complete cessation of sperm development. However, the mechanisms of interaction between T. gondii and the reproductive system is unclear. The aims were to study the expression levels of genes related to spermatogenesis, following T. gondii infection, in mouse testicular tissue.
METHODS: RNA-seq sequencing was carried out on mouse testicular tissues from mice infected or uninfected with the T. gondii type II Prugniaud (PRU) strain and validated in combination with real-time quantitative PCR and immunofluorescence assays.
RESULTS: The results showed that there were 250 significant differentially expressed genes (DEGs) (P < 0.05, |log2fold change| ≧ 1). Bioinformatics analysis showed that 101 DEGs were annotated to the 1696 gene ontology (GO) term. While there was a higher number of DEGs in the biological process classification as a whole, the GO enrichment revealed a significant presence of DEGs in the cellular component classification. The Arhgap18 and Syne1 genes undergo regulatory changes following T. gondii infection, and both were involved in shaping the cytoskeleton of the blood-testis barrier (BTB). The number of DEGs enriched in the MAPK signaling pathway, the ERK1/2 signaling pathway, and the JNK signaling pathway were significant. The PTGDS gene is located in the Arachidonic acid metabolism pathway, which plays an important role in the formation and maintenance of BTB in the testis. The expression of PTGDS is downregulated subsequent to T. gondii infection, potentially exerting deleterious effects on the integrity of the BTB and the spermatogenic microenvironment within the testes.
CONCLUSIONS: Overall, our research provides in-depth insights into how chronic T. gondii infection might affect testicular tissue and potentially impact male fertility. These findings offer a new perspective on the impact of T. gondii infection on the male reproductive system.

To learn about the gene structure, phylogenetic evolution, and function under biotic and abiotic stresses of BTB (Bric-a-Brac/Tramtrack/Broad Complex) genes in Paulownia fortunei, a whole-genome sequence evaluation was carried out, and a total of 62 PfBTB genes were identified. The phylogenetic analysis showed that PfBTB proteins are divided into eight groups, and these proteins are highly conserved. PfBTB genes were unevenly distributed on 17 chromosomes. The colinearity analysis found that fragment replication and tandem replication are the main modes of gene amplification in the PfBTB family. The analysis of cis-acting elements suggests that PfBTB genes may be involved in a variety of biological processes. The transcriptomic analysis results showed that PfBTB3/12/14/16/19/36/44 responded to Paulownia witches\' broom (PaWB), while PfBTB1/4/17/43 responded to drought stress, and the RT-qPCR results further support the reliability of transcriptome data. In addition, the association analysis between miRNA and transcriptome revealed a 91-pair targeting relationship between miRNAs and PfBTBs. In conclusion, the BTB genes in Paulownia are systematically identified in this research. This work provides useful knowledge to more fully appreciate the potential functions of these genes and their possible roles in the occurrence of PaWB and in response to stress.

We have previously reported that serum albumin-coated bone allograft (BoneAlbumin, BA) is an effective bone substitute. It improves bone regeneration at the patellar and tibial donor sites six months after harvesting bone-patellar tendon-bone (BPTB) autografts for primary anterior cruciate ligament reconstruction (ACLR). In the present study, we examined these donor sites seven years after implantation. The study group (N = 10) received BA-enhanced autologous cancellous bone at the tibial and BA alone at the patellar site. The control group (N = 16) received autologous cancellous bone at the tibial and blood clot at the patellar site. We evaluated subcortical density, cortical thickness, and bone defect volume via CT scans. At the patellar site, subcortical density was significantly higher in the BA group at both time points. There was no significant difference in cortical thickness between the two groups at either donor site. The control group\'s bone defect significantly improved and reached the BA group\'s values at both sites by year seven. Meanwhile, the bone defects in the BA group did not change significantly and were comparable to the six-month measurements. No complications were observed. There are two limitations in this study: The number of patients recruited is small, and the randomization of the patients could have improved the quality of the study as the control group patients were older compared to the study group patients. Our 7-year results seem to demonstrate that BA is a safe and effective bone substitute that supports faster regeneration of donor sites and results in good-quality bone tissue at the time of ACLR with BPTB autografts. However, studies with a larger number of patients are required to definitively confirm the preliminary results of our study.

KEAP1 promotes the ubiquitin-dependent degradation of NRF2 by assembling into a CUL3-dependent ubiquitin ligase complex. Oxidative and electrophilic stress inhibit KEAP1 allowing NRF2 to accumulate for the transactivation of stress response genes. To date there are no structures of the KEAP1-CUL3 interaction nor binding data to show the contributions of different domains to their binding affinity. We determined a crystal structure of the BTB and 3-box domains of human KEAP1 in complex with the CUL3 N-terminal domain that showed a heterotetrameric assembly with 2:2 stoichiometry. To support the structural data, we developed a versatile TR-FRET-based assay system to profile the binding of BTB-domain-containing proteins to CUL3 and determine the contribution of distinct protein features, revealing the importance of the CUL3 N-terminal extension for high affinity binding. We further provide direct evidence that the investigational drug CDDO does not disrupt the KEAP1-CUL3 interaction, even at high concentrations, but reduces the affinity of KEAP1-CUL3 binding. The TR-FRET-based assay system offers a generalizable platform for profiling this protein class and may form a suitable screening platform for ligands that disrupt these interactions by targeting the BTB or 3-box domains to block E3 ligase function.

UNASSIGNED: The primary goals of this study were to investigate the potential roles of ZNF22 and HDAC3 as a histone deacetylase in regulating an increases in blood-tumor barrier (BTB) permeability and some of the possible molecular mechanisms associated with this effect.
UNASSIGNED: The expression of ZNF22 and HDAC3 in glioma-exposed endothelial cells (GECs) of BTB were detected transcription real-time PCR or western blot. The interaction of ZNF22 and HDAC3 in GECs associated with transcript effect was analyzed by means of Co-Immunoprecipitation and luciferase reporter assay.
UNASSIGNED: In the present investigation, GECs expressed higher levels of ZNF22 as a zinc finger transcription factor and HDAC3 than endothelial cells. We then affirmed that silencing HDAC3 or ZNF22 led to a reduction in BTB permeability. By bioinformatics analysis, chromatin immunoprecipitation (ChIP) assays and luciferase assay, we found that ZNF22 had a target binding relationship with the promoter regions of ZO-1, Occludin, and Claudin-5 and negatively regulated the expression of ZO-1, Occludin, and Claudin-5. Furthermore, we revealed that HDAC3, as a co-transcript repressor with histone deacetylase activity, could interact with ZNF22 to hinder the expression of TJ-associated proteins, thereby further facilitating the permeability of BTB.
UNASSIGNED: ZNF22 acted as a transcription factor in conjunction with HDAC3 to modulate the expression of TJ-associated proteins, which was correlated with an increase in BTB permeability. These results may provide new strategies and targets for the chemotherapy of gliomas as well as intracranial infections.

Microtubule affinity-regulating kinases (MARKs) are nonreceptor Ser/Thr protein kinases known to regulate cell polarity and microtubule dynamics in Caenorhabditis elegans, Drosophila, invertebrates, vertebrates, and mammals. An earlier study has shown that MARK4 is present at the ectoplasmic specialization and blood-testis barrier (BTB) in the seminiferous epithelium of adult rat testes. Here, we report the function of MARK4 and another isoform MARK2 in Sertoli cells at the BTB. Knockdown of MARK2, MARK4, or MARK2 and MARK4 by RNAi using the corresponding siRNA duplexes without apparent off-target effects was shown to impair tight junction (TJ)-permeability barrier at the Sertoli cell BTB. It also disrupted microtubule (MT)- and actin-based cytoskeletal organization within Sertoli cells. Although MARK2 and MARK4 were shown to share sequence homology, they likely regulated the Sertoli cell BTB and MT cytoskeleton differently. Disruption of the TJ-permeability barrier following knockdown of MARK4 was considerably more severe than loss of MARK2, though both perturbed the barrier. Similarly, loss of MARK2 affected MT organization in a different manner than the loss of MARK4. Knockdown of MARK2 caused MT bundles to be arranged around the cell periphery, whereas knockdown of MARK4 caused MTs to retract from the cell edge. These differences in effects on the TJ-permeability barrier are likely from the unique roles of MARK2 and MARK4 in regulating the MT cytoskeleton of the Sertoli cell.

Male reproductive function is key to the continuation of species and is under sophisticated regulation, challenged by various stressors including inflammation. In the lipopolysaccharide (LPS) intraperitoneal injection-induced acute systemic inflammation, male fecundity was compromised with decreased testosterone level, damaged spermatogenesis, and downregulations of testicular gene expression levels involved in steroidogenesis regulation and blood-testis barrier. It is also noteworthy that the testis is more sensitive to acute stress caused by LPS-induced systemic inflammation. LPS treatment resulted in lower testicular gene expression levels of steroidogenic acute regulatory protein, cholesterol side-chain cleavage enzyme, and cytochrome P450 family 11 subfamily B member 1 after LPS treatment, while no such decrease was found in the adrenal gland. In parallel to the significant decreases in testicular intercellular adhesion molecule 1, tight junction protein 1, and gap junction alpha-1 protein gene expression with LPS treatment, no decrease was found in the epididymis. In the brain, LPS treatment caused higher medial preoptic area (mPOA) activation in the hypothalamus, which is accompanied by elevated blood follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels, suggesting a disturbed hypothalamic-pituitary-gonad axis function. Besides mPOA, brain c-fos mapping and quantitative analysis demonstrated a broad activation of brain nuclei by LPS, including the anterior cingulate cortex, lateral septum, paraventricular nucleus of the hypothalamus, basolateral amygdala, ventral tegmental area, lateral habenular nucleus, locus coeruleus, Barrington\'s nucleus, and the nucleus of the solitary tract, accompanied by abnormal animal behavior. Our data showed that LPS-induced inflammation caused not only local testicular damage but also a systemic disturbance at the brain-testis axis level.

BACKGROUND: Bovine tuberculosis is an important zoonotic disease caused by Mycobacterium bovis, known to exist in all parts of the world especially in un-hygienic environments. It has a public health impact and humans acquire the infection from infected animals through consumption of raw or unpasteurized milk and dairy products.
METHODS: An Institutional-based quantitative cross-sectional study was conducted from February to April 2020 to assess the knowledge and practice level along with their derivatives of bovine tuberculosis among adult HIV-positive people in Bahir Dar city public hospitals. The data were collected using structured interviewer questionnaire and systematic random sampling was employed to select study subjects. Thus, a total of 435 people living with HIV were considered for this study.
RESULTS: Out of the total 435 participants 51.5% were male and the mean age was 37.6 years. The overall level of good knowledge and practice of HIV-positive persons for bovine TB were 46.2% and 41.1%, respectively. Moreover, education status, religion, cattle holding, and environmental hygiene were statistically significant associations (p < 0.05) with knowledge level. Likewise, education status, age, and marital status were statistically significant associations with the practice level for bovine TB. Education was a key determinant factor affecting both knowledge and practice.
CONCLUSIONS: In conclusion, the present study showed that the level of good knowledge and practice of the participant for bovine TB was low. Hence, tailor-made training by medical and veterinary professionals on the identified risk factors is recommended.

It has been found that polystyrene microplastics (PS-MPs) exposure leads to decreased sperm quality and quantity, and we aim to explore the underlying mechanisms. Therefore, we gave 20 mg/kg body weight (bw) and 40 mg/kg bw 4 μm and 10 μm PS-MPs to male Balb/c mice by gavage. RNA sequencing of testes was performed. After PS-MPs exposure, blood-testis barrier (BTB) integrity was impaired. Since cytoskeleton was closely related to BTB integrity maintenance, and cytoskeleton disorganization could be induced by PS-MPs exposure in the testis, which resulted in the truncation of actin filaments and disruption of BTB integrity. Such processes were attributed to the differential expression of Arp3 and Eps8 (two of the most important actin-binding proteins). According to the transcriptome sequencing results, we examined the oxidative stress level in the testes and Sertoli cells. We found that PS-MPs exposure induced increased reactive oxygen species (ROS) level, which destroyed the balance between mTORC1 and mTORC2 (the mTORC1 activity was increased, while the mTORC2 activity was decreased). In conclusion, PS-MPs induced the imbalance of mTORC1 and mTORC2 via the ROS burst, and altered the expression profile of actin-binding proteins, resulting in F-actin disorganization and reduced expression of junctional proteins in the BTB. Eventually PS-MPs led to BTB integrity disruption and spermatogenesis dysfunction.

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