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Impaired spermatogenesis and male infertility are common manifestations associated with mitochondrial diseases, yet the underlying mechanisms linking these conditions remain elusive. In this study, we demonstrate that mice deficient for the mitochondrial intra-membrane rhomboid protease PARL, a recently reported model of the mitochondrial encephalopathy Leigh syndrome, develop early testicular atrophy caused by a complete arrest of spermatogenesis during meiotic prophase I, followed by degeneration and death of arrested spermatocytes. This process is independent of neurodegeneration. Interestingly, genetic modifications of PINK1, PGAM5, and TTC19 – three major substrates of PARL with important roles in mitochondrial homeostasis – fail to reproduce or modify this severe phenotype, indicating that the spermatogenic arrest arises from distinct molecular pathways. We further observed severe abnormalities in mitochondrial ultrastructure in PARL-deficient spermatocytes, along with prominent electron transfer chain defects, disrupted coenzyme Q (CoQ) biosynthesis, and metabolic rewiring. These mitochondrial defects are associated with a germ cell-specific decrease in GPX4 expression leading arrested spermatocytes to ferroptosis – a regulated cell death modality characterized by uncontrolled lipid peroxidation. Our results suggest that mitochondrial defects induced by PARL depletion act as an initiating trigger for ferroptosis in primary spermatocytes through simultaneous effects on GPX4 and CoQ – two major inhibitors of ferroptosis. These findings shed new light on the potential role of ferroptosis in the pathogenesis of mitochondrial diseases and male infertility warranting further investigation. Up to 9% of men are thought to experience infertility. These individuals may not produce enough healthy sperm cells. The root cause of infertility is often not discovered but, in some cases, it is associated with genetic defects in cell compartments known as mitochondria. Mitochondria are responsible for converting energy from food into a form of chemical energy cells need to power vital processes. However, it remains unclear how defects in mitochondria contribute to male infertility. Leigh syndrome is one of the most prevalent and severe diseases caused by genetic defects in mitochondria. The condition often develops in childhood and affects the nervous system, muscle and other organs, leading to many symptoms including muscle weakness and neurological regression. A previous study found that mutant mice that lack an enzyme, called PARL, display symptoms that are similar to those observed in humans with Leigh syndrome. PARL is found inside mitochondria where it cuts specific proteins to ensure they are working correctly in the cells. Radaelli et al. used extensive microscopy and biochemical analyses to study the fertility of male mice lacking PARL. The experiments revealed that the males were infertile due to a failure to produce sperm: spermatocytes, which usually develop into sperm cells, where much more likely to die in mice without PARL (by a process known as ferroptosis). Further experiments demonstrated that the mitochondria of the mutant mice had a shortage of two crucial molecules, a protein called GPX4 and a lipid called Coenzyme Q, which are required to prevent death by ferroptosis. It appears that this shortage was responsible for the demise of spermatocytes in the male mutant mice affected by infertility. These findings reveal a new role for PARL in the body and provide evidence that mitochondrial defects in living mammals can trigger ferroptosis, thereby contributing to male infertility. In the future, this research may pave the way for new treatments for male infertility and other diseases associated with defects in mitochondria.

Inherited retinal diseases (IRDs) are a diverse group of disorders that share common vision deficits ranging from early onset blindness to severe and progressive later-onset disease. We report a form of early-onset day-vision loss, cone-rod dystrophy, in the Standard poodle. Through GWAS and homozygosity mapping, a large deletion on CFA8:NC_049229.1:g.60,022,583_60,040,453del was found which removes 3' portions of two different genes, PTPN21 and SPATA7, presenting a challenge for assessing the actual causative gene in a multi-gene large deletion. All affected dogs were homozygous for the mutant allele, which segregated perfectly with the phenotype within the breed. The variant was absent in 1879 dogs from the Dog10K database. While the role of SPATA7 for retinal disease has been established in human patients and genetically engineered mice, the role of PTPN21 in the retina is unclear even though it is expressed in rod and cone photoreceptors. Expression of whole and truncated transcripts for both genes was detected in skin fibroblasts from controls and cases. Retinal RNA analysis of PTPN21 splicing suggests that at least one unmodified transcript is still present in mutants. Ptpn21-/- knockout mice did not have an ocular phenotype, and IHC for rod- and cone-specific opsins detected no cone or rod abnormalities suggesting that PTPN21 loss has minimal to no contributory role towards the retinal phenotype in mutants. The variant leads to a deletion of the 3'-end of the SPATA7 transcript: XM_038545497.1:r.1,314_1,629delins[g.60,018,954-60,018,990], p.(XP_038401425.1: Asp361GlufsTer2), reducing the predicted protein from 595 to 361 AA. Ultrastructure expansion microscopy (U-ExM) enabled the detection of a distinct SPATA7 signal around the transition zone of the primary cilium in photoreceptors and fibroblasts of WT dogs, which was absent in affected dog. We posit that SPATA7 deficiency is the main cause of the condition, and propose this disease as a model for the SPATA7-related form of cone-rod dystrophy in humans. Our work shows an example of functional refinement of a multi-gene deletion variant using a multi-technique approach.

BackgroundEngraftment efficiency of human immune cell lineages in super immunodeficient mice varies, and numerous strains have been developed to improve reconstitution of humanized immune system mice. NSG-SGM3 and NOG-EXL mice combine severe immunodeficiency with transgenic expression of human myeloid stimulatory cytokines, resulting in improved expansion of myeloid populations. However, humanized NSG-SGM3 (huNSG-SGM3) mice develop a lethal macrophage activation syndrome (MAS) and mast cell hyperplasia that limit their use in long-term studies, especially those requiring humanization followed by tumor xenotransplantation. It is currently unclear to what extent humanized NOG-EXL (huNOG-EXL) mice suffer from the same conditions observed in huNSG-SGM3 mice. In this study, we aimed to compare the effects of human CD34+ hematopoietic stem cell engraftment in these two mouse strains in an orthotopic glioblastoma patient-derived organoid xenograft model.MethodsNSG-SGM3 mice (n=10) humanized in-house were compared to both NOG-EXL mice (n=10) humanized in-house and to commercially available huNOG-EXL mice (n=12). Mice were euthanized at humane or study endpoints, and a complete pathological assessment was performed. A semiquantitative multiparametric clinicopathological scoring system was developed to characterize the myeloid proliferative disorder.ResultsBoth the NOG-EXL mice humanized in-house and commercially available huNOG-EXL mice survived longer (to experimental endpoint) than huNSG-SGM3 mice (22 vs 17 weeks post engraftment), with significantly less severe MAS and lack of mastocytic proliferation. Major findings included mast cell infiltration of the pancreas and liver (huNSG-SGM3 only), increased eosinophilopoiesis, anemia, and histiocytic infiltration of the spleen (both strains). Engraftment of human lymphocytes, assessed by immunohistochemistry, was similar in the two strains. The longer survival and decreased MAS severity in NOG-EXL mice enabled their use in a patient-derived xenograft transplantation study.ConclusionsHumanized NOG-EXL mice develop a milder myeloid activation syndrome and do not develop mast cell hyperplasia relative to humanized NSG-SGM3 mice. Thus, the NOG-EXL model may be better suited than the NSG-SGM3 model for immuno-oncology studies requiring long-term survival post humanization. Future directions include further assessment of the lymphocyte populations in both strains and application of the humanized NOG-EXL for patient-derived xenograft models.AcknowledgementsThe authors thank the staff of the Penn Vet Histology Lab and the Penn Stem Cell and Xenograft Core for technical assistance.Ethics ApprovalAll the mice included in this study were maintained by the Stem Cell and Xenograft Core at the University of Pennsylvania under the protocol #803506 which has been reviewed and approved by the Institutional Animal Care and Use Committee (IACUC). The participant gave informed consent for use of glioblastoma patient-derived organoids, under protocol #832366, approved by the University of Pennsylvania Institutional Review Board (IRB).

Studies from the emerging field of Positive Psychology hypothesise that personal resources like the constructs of Psychological Capital (i.e. Hope, Optimism, Self-Efficacy and Resilience) may contribute to decreased stress and increased work engagement. The present study aims to explore the relationship between employee engagement, perceived stress and psychological capital of employees working in private sector organisation in terms of gender difference and duration of service. A General Information Schedule, the Utrecht Work Engagement Scale, Perceived Stress Scale and Psychological Capital Questionnaire were used as tools. Both qualitative and quantitative analyses were done. The findings revealed that psychological capital among the male employees is comparatively higher than that of the female employees. On the contrary, perceived stress is comparatively higher among the female employees compared to the male employees. The findings also revealed that the more the duration of service, the more is the employee engagement. Employee engagement and psychological capital are positively correlated. On the other hand, perceived stress was negatively correlated with employee engagement and also with psychological capital. Measures may be taken to improve psychological capital and engagement and curb stress among employees to foster a positive working environment.

The normal tissue sparing afforded by FLASH radiotherapy (RT) is being intensely investigated for potential clinical translation. Here, we studied the effects of FLASH proton RT (F-PRT) in the reirradiation setting, with or without hypofractionation. Chronic toxicities in three murine models of normal tissue toxicity including the intestine, skin, and bone were investigated. In studies of the intestine, single-dose irradiation was performed with 12 Gy of Standard proton RT (S-PRT), followed by a second dose of 12 Gy of F-PRT or S-PRT. Additionally, a hypofractionation scheme was applied in the reirradiation setting (3 x 6.4 Gy of F-PRT or S-PRT, given every 48 hrs). In studies of skin/bone of the murine leg, 15 Gy of S-PRT was followed by hypofractionated reirradiation with F-PRT or S-PRT (3 x 11 Gy). Compared to reirradiation with S-PRT, F-PRT reduced intestinal fibrosis and collagen deposition in the reirradiation setting and significantly increased survival rate, demonstrating its protective effects on intestinal tissues. In previously irradiated leg tissues, reirradiation with hypofractionated F-PRT created transient dermatitis that fully resolved in contrast to reirradiation with hypofractionated S-PRT. Lymphedema was also alleviated after a second course of radiation with F-PRT, along with significant reductions in the accumulation of fibrous connective tissue in the skin compared to mice reirradiated with S-PRT. The delivery of a second course of fractionated S-PRT induced tibial fractures in 83.3% of the mice, whereas only 20% of mice reirradiated with F-PRT presented with fractures. These studies provide the first evidence of the sparing effects of F-PRT, in the setting of hypofractionated reirradiation. The results support FLASH as highly relevant to the reirradiation regimen where it exhibits significant potential to minimize chronic complications for patients undergoing RT.

This study aims to develop a semi-automatically labelled prosody database for Hindi, for enhancing the intonation component in ASR and TTS systems, which is also helpful for building Speech to Speech Machine Translation systems. Although no single standard for prosody labelling exists in Hindi, researchers in the past have employed perceptual and statistical methods in literature to draw inferences about the behaviour of prosody patterns in Hindi. Based on such existing research and largely agreed upon theories of intonation in Hindi, this study attempts to first develop a manually annotated prosodic corpus of Hindi speech data, which is then used for training prediction models for generating automatic prosodic labels. A total of 5,000 sentences (23,500 words) for declarative and interrogative types have been labelled. The accuracy of the trained models for pitch accent, intermediate phrase boundaries and accentual phrase boundaries is 73.40%, 93.20%, and 43% respectively.

Impaired spermatogenesis and male infertility are common manifestations of mitochondrial diseases, but the underlying mechanisms are unclear. Here we show that mice deficient for PARL, the mitochondrial rhomboid protease, a recently reported model of Leigh syndrome, develop postpubertal testicular atrophy caused by arrested spermatogenesis and germ cell death independently of neurodegeneration. Genetic modifications of PINK1, PGAM5, and TTC19, three major substrates of PARL with important roles in mitochondrial homeostasis, do not reproduce or modify this phenotype. PARL deficiency in testis mitochondria leads to severe mitochondrial electron transfer chain defects, alterations in Coenzyme Q biosynthesis and redox status, and abrogates GPX4 expression specifically in spermatocytes leading to massive ferroptosis, an iron-dependent regulated cell death modality characterized by uncontrolled lipid peroxidation. Thus, mitochondrial defects can initiate ferroptosis in vivo in specific cell types by simultaneous effects on GPX4 and Coenzyme Q. These results highlight the importance of ferroptosis and cell-type specific downstream responses to mitochondrial deficits with respect to specific manifestations of mitochondrial diseases.Competing Interest StatementThe authors have declared no competing interest.

Background: Community-acquired pneumonia (CAP) remains one of the leading causes of mortality and morbidity in children under 5 years of age. Early diagnosis and effective management remain the cornerstone to reduce complications and fatalities associated with CAP. Lately, C-reactive protein (CRP) and procalcitonin (PCT) have gained much attention as useful biomarkers in CAP. Recent data suggest that a very low PCT value has a high negative predictive value while identifying a population of children at low risk of typical bacterial infections. Hence, we propose to take up this study to assess the clinical utility of PCT in pediatric CAP. Aims and Objectives: The aim of this study was to study serum PCT level in children with the clinical diagnosis of CAP admitted at R.G Kar Medical College and Hospital, Kolkata, and its role in severity identification and contribution in the treatment of CAP. Materials and Methods: This cross-sectional hospital-based observational study was performed from January 2020 to June 2021 in the R.G Kar Medical College and Hospital, Kolkata, West Bengal, India. A total of 50 children were included within the age group of 2 months–5 years with CAP admitted in the Children’s ward and intensive care unit at our hospital. Clinical characteristics such as age, sex, symptoms such as history of fever, difficulty feeding or drinking, convulsion, lethargy, cyanosis, chest in-drawing, respiratory rates, auscultatory findings such as air entry of bilateral lungs, added sounds like-(inspiratory crept, bronchial breathing), and oxygen saturation were studied. Routine investigations such as complete blood count, serum CRP, blood culture, and chest radiograph sent along with serum PCT. Results: Out of 50 children diagnosed with CAP, 30 (60%) fell within the age range of 2–5 months of age. Among the study population, 70% of children displayed weakly positive and 20% strongly positive serum PCT values. A strong correlation (P < 0.001) was revealed between the severity of pneumonia and PCT, and also a positive correlation (P < 0.001) between clinical outcome and PCT. However, the role in contribution in early diagnosis could not be concluded. Conclusion: This study concluded that serum PCT may be used as a valuable indicator for assessing the severity and prognosis of CAP. All children with elevated PCT who were prescribed antibiotics did not experience further complications of CAP.