Delineating the Disease Boundaries: Homozygous CDC14A Variants Underlying Nonsyndromic Hearing Loss and Hearing Impairment Infertile Male Syndrome.

Background: Hereditary hearing loss is a genetically heterogeneous neurosensory disorder that affects many people. Deafness and infertility can coexist in some cases, creating the hearing impairment infertile male syndrome. There are several known molecular mechanisms that can cause deafness either on its own or in conjunction with infertility. Methods and Results: Here, we represent two consanguineous families (A, B), both families had clinical evidence of deafness, and family B also had infertility, so we referred to them as having nonsyndromic hearing loss (NSHL) and hearing impairment infertile male syndrome (HIIMS), respectively. These families' genetic makeup was examined using an Affymetrix GeneChip 250K Nsp array followed by Sanger sequencing. In family A, we identified a novel homozygous stop gain variant [NM_003672.4; c.1000C>T; p.(Gln334*)] and a homozygous missense variant [NM_003672.4; c.684C>A; p.(Asn228Lys)] in family B in CDC14A gene (MIM#603504). In animal models, the CDC14A gene causes both hearing loss and infertility; in addition, it also causes NSHL and HIIMS in humans. Conclusions: Our study on the CDC14A gene has identified two novel variants, crucial for delineating disease boundaries. Variants in exon 10 and upstream cause HIIMS, and those in exon 11 and downstream are linked exclusively to hearing impairment. This precision enhances diagnostics and offers potential for targeted interventions, marking a significant advancement in understanding the genetic basis of these conditions.

FSP4HSP: Frequent sequential patterns for the improved classification of heat shock proteins, their families, and sub-types.

Heat shock proteins (HSPs) from different families and sub-types play a vital role in the folding and unfolding of proteins, in maintaining cellular health, and in preventing serious disorders. Previous computational methods for HSP classification have yielded promising performance. However, most of the existing methods rely heavily on amino acid composition features and still face challenges related to interpretability and accuracy. To overcome these issues, we introduce a novel frequent sequential pattern (FSP)-based analysis and classification method for the classification of HSPs, their families, and sub-types. The proposed method is called FSP4HSP, which stands for "FSP for HSP". It identifies FSPs of amino acids (FSPAAs) and utilizes them for analysis and classification. Besides FSPAAs, sequential rules among amino acids are also discovered. Both binary and multi-class classification scenarios are considered, with the utilization of eight integer-based and four string-based classifiers. The incorporation of FSPAAs in the classification/prediction task enhances the interpretability of FSP4HSP and a comprehensive performance comparison using various evaluation measures demonstrates that it surpasses existing methods for the classification/recognition of HSPs.

Clinical and genetic investigation of 14 families with various forms of short stature syndromes.

Skeletal dysplasias are a heterogeneous group of disorders presenting mild to lethal defects. Several factors, such as genetic, prenatal, and postnatal environmental may contribute to reduced growth. Fourteen families of Pakistani origin, presenting the syndromic form of short stature either in the autosomal recessive or autosomal dominant manner were clinically and genetically investigated to uncover the underlying genetic etiology. Homozygosity mapping, whole exome sequencing, and Sanger sequencing were used to search for the disease-causing gene variants. In total, we have identified 13 sequence variants in 10 different genes. The variants in the HSPG2 and XRCC4 genes were not reported previously in the Pakistani population. This study will expand the mutation spectrum of the identified genes and will help in improved diagnosis of the syndromic form of short stature in the local population.

SPM4GAC: SPM based approach for genome analysis and classification of macromolecules.

Genome sequence analysis and classification play critical roles in properly understanding an organism's main characteristics, functionalities, and changing (evolving) nature. However, the rapid expansion of genomic data makes genome sequence analysis and classification a challenging task due to the high computational requirements, proper management, and understanding of genomic data. Recently proposed models yielded promising results for the task of genome sequence classification. Nevertheless, these models often ignore the sequential nature of nucleotides, which is crucial for revealing their underlying structure and function. To address this limitation, we present SPM4GAC, a sequential pattern mining (SPM)-based framework to analyze and classify the macromolecule genome sequences of viruses. First, a large dataset containing the genome sequences of various RNA viruses is developed and transformed into a suitable format. On the transformed dataset, algorithms for SPM are used to identify frequent sequential patterns of nucleotide bases. The obtained frequent sequential patterns of bases are then used as features to classify different viruses. Ten classifiers are employed, and their performance is assessed by using several evaluation measures. Finally, a performance comparison of SPM4GAC with state-of-the-art methods for genome sequence classification/detection reveals that SPM4GAC performs better than those methods.

A prospective observational study of estimating drug related problems and clinical outcomes in subtypes of stroke patients.

BACKGROUND: Stroke is a neurological disease and a leading cause of mortality worldwide. Strokes mainly consist of two types: hemorrhage and ischemia. Stroke patients are being administered multiple drug therapy and are at risk of drug-related problems. AIM: To estimate drug-related problems (DRPs) and clinical end outcomes in hospitalized stroke patients. METHODS: Current study was a multicenter, cross-sectional prospective observational study including 250 stroke patients admitted to tertiary care hospitals in Karachi, Pakistan. The study included all clinical subtypes of stroke patients i.e. Stroke, Ischemic stroke, Hemorrhagic stroke, CVA, and TIA. Associations among patient-clinical end outcomes and drug therapy-related variables like DRPs, mortality, and morbidity rates were estimated using Pearson's chi-squared test. Statistical analysis was done by using SPSS software, version 25. RESULTS: A total of 250 patients participated in this study suffering from different clinical subtypes of stroke i.e. Ischemic stroke, hemorrhagic stroke, TIA, and CVA, including 46% male and 54% female patients. The majority of patients' stay at the hospital was between 1-10 days. The overall mortality rate in stroke patients was 51%. HAIs were observed in 70% of patients, HAIs faced by patients were SAP, CAP, UTI, sepsis, and VAP. Drugs were assessed according to NEML i.e. access group antibiotics, watch group antibiotics, reserve group antibiotics, statins, antiepileptics, and proton pump inhibitors. Majorly ceftriaxone was administered to 79% of patients, piperacillin-tazobactam to 52%, and cefixime to 48%, whereas meropenem was administered to 42% of patients along with vancomycin to 39% of total patients. A high mortality rate was observed in the case of Klebsiella pneumoniae and Staphylococcus aureus i.e. 78% and in the case of streptococcus pneumoniae 61% mortality rate was observed. Due to the presence of DRPs and various other clinical factors like comorbidities, DDIs, HAIs, administration of potentially nephrotoxic drugs, and administration of antibiotics without having CST, hospitalized stroke patients faced many problems. CONCLUSION: This study helped determine DRPs along with various clinical factors affecting the clinical end outcomes of patients suffering from any clinical subtype of stroke. Due to the enhancement in the evidence of the incidence of DRPs in tertiary care hospitals, pharmacist-led drug therapy review by interfering with doctors and other medical professionals at the patient bed site is needed and should be done to avoid any negative end outcomes and serious issues related to DRPs.

A variant in sperm-specific glycolytic enzyme enolase 4 (ENO4) causes human male infertility.

BACKGROUND: Although defects in sperm morphology and physiology lead to male infertility, in many instances, the exact disruption of molecular pathways in a given patient is often unknown. The glycolytic pathway is an essential process to supply energy in sperm cell motility. Enolase 4 (ENO4) is crucial for the glycolytic process, which provides the energy for sperm cells in motility. ENO4 is located in the sperm principal piece and is essential for the motility and organization of the sperm flagellum. In the present study, we characterized a family with asthenozoospermia and abnormal sperm morphology as a result of a variant in the enolase 4 (ENO4) gene. METHODS: Computer-assisted semen analysis, papanicolaou smear staining and scanning electron microscopy were used to examine sperm motility and morphology for semen analysis in patients. For genetic analysis, whole-exome sequencing followed by Sanger sequencing was performed. RESULTS: Two brothers in a consanguineous family were being clinically investigated for sperm motility and morphology issues. Genetic analysis by whole-exome sequencing revealed a homozygous variant [c.293A>G, p.(Lys98Arg)] in the ENO4 gene that segregated with infertility in the family, shared by affected but not controls. CONCLUSIONS: In view of the association of asthenozoospermia and abnormal sperm morphology in Eno4 knockout mice, we consider this to be the first report describing the involvement of ENO4 gene in human male infertility. We also explore the possible involvement of another variant in explaining other phenotypic features in this family.

LRRC23 truncation impairs radial spoke 3 head assembly and sperm motility underlying male infertility.

Radial spokes (RS) are T-shaped multiprotein complexes on the axonemal microtubules. Repeated RS1, RS2, and RS3 couple the central pair to modulate ciliary and flagellar motility. Despite the cell type specificity of RS3 substructures, their molecular components remain largely unknown. Here, we report that a leucine-rich repeat-containing protein, LRRC23, is an RS3 head component essential for its head assembly and flagellar motility in mammalian spermatozoa. From infertile male patients with defective sperm motility, we identified a splice site variant of LRRC23. A mutant mouse model mimicking this variant produces a truncated LRRC23 at the C-terminus that fails to localize to the sperm tail, causing male infertility due to defective sperm motility. LRRC23 was previously proposed to be an ortholog of the RS stalk protein RSP15. However, we found that purified recombinant LRRC23 interacts with an RS head protein RSPH9, which is abolished by the C-terminal truncation. Evolutionary and structural comparison also shows that LRRC34, not LRRC23, is the RSP15 ortholog. Cryo-electron tomography clearly revealed that the absence of the RS3 head and the sperm-specific RS2-RS3 bridge structure in LRRC23 mutant spermatozoa. Our study provides new insights into the structure and function of RS3 in mammalian spermatozoa and the molecular pathogenicity of LRRC23 underlying reduced sperm motility in infertile human males.

Attenuation of CFA-induced chronic inflammation by a bicyclic monoterpene fenchone targeting inducible nitric oxide, prostaglandins, C-reactive protein and urea.

Fenchone (a bicyclic monoterpene) is present in the essential oils of plant species like Foeniculum vulgare and Peumus boldus and is used to treat GIT disorders. Research reports have indicated its strong anti-inflammatory, antioxidant, and anti-nociceptive properties. The present study was designed to investigate fenchone's anti-arthritic effects in a rat model of chronic joint inflammation (Complete Freud's Adjuvant-mediated inflammation [CFA]). Molecular docking analysis revealed a high binding interaction of fenchone with inducible nitric oxide synthase (iNOS), Interleukin-17, Prostaglandin E Receptor EP4, and Cycloxygenase-2 (COX-2), indicating its anti-inflammatory efficacy using computational tests. Fenchone treatment at 100 mg/kg, 200 mg/kg, and 400 mg/kg significantly enhanced the tail-flick latency when compared with the solvent-treated group. Correspondingly, the raised mRNA values of iNOS, IL-17, IL-1ß, IL-6, TNF-α, and COX-2 in solvent-treated group were significantly reduced following treatment with fenchone. Moreover, fenchone significantly lowered spleen and thymus indices, Nitric oxide (NO) and PGE2 values as compared to solvent-treated group. Hence, the results of the present study indicated that fenchone has a potent anti-inflammatory effect by inhibiting pro-inflammatory markers and thus may have therapeutic potential for chronic joint inflammation as well as chronic inflammatory disorders.

Linalool: Monoterpene alcohol effectiveness in chronic synovitis through lowering Interleukin-17, spleen and thymus indices.

Interleukin-17 has a positive role in the initial induction and late chronic phases of many inflammatory disorders like arthritis. This cytokine has a strong option for therapeutic targeting due to the fact that it was found in the inflamed joints of individual with rheumatoid arthritis (RA) and persuasive evidence from experimental arthritis models indicating its pro-inflammatory actions. IL-17 suppression lessened the asperity of arthritis. The present study aimed to assess the anti-arthritic potential of linalool in a model of chronic joint inflammation (CFA-mediated rheumatoid arthritis) in rats. Linalool markedly lowered spleen and thymus indices as opposed to arthritic control. The over-formation of IL-17, COX-2, TNF-α IL-1ß, iNOS and IL-6 were markedly impaired in all linalool treated rats, but IL-10 was raised as compared to arthritic animals in Real time-PCR. There was reduction in associated parameters like paw volume, arthritic index, mobility score, and flexion pain score and a marked increase in stance score in CFA model as compared to the arthritic control group. Furthermore, there was improvement in body weight, hematological, tissue, and radiological parameters in the CFA-model. Molecular docking study exhibited strong binding interaction of linalool with IL-17, PGE-2, iNOS and COX-2, thus providing a good correlation among experimental and theoretical results. The current findings show that linalool reduces adjuvant arthritis by suppressing pro-inflammatory mediators, arthritic development, and spleen and thymus indices. Thus, linalool may be employed therapeutically to alleviate arthritis in humans.

Asymptomatic COVID-19 presenting with features of mixed pattern acute liver injury in a young healthy female, a case report.

COVID-19 associated severe acute liver injury in a young healthy patient has not been reported much in the literature. And currently, there are no standard management guidelines. We want to report a case of acute liver injury of mixed pattern in a young healthy female with asymptomatic COVID-19 infection. She presented with abdominal pain, nausea, vomiting and yellowish discoloration of her skin. Further laboratory investigations revealed mixed pattern liver injury with highly raised liver enzymes. She was managed with N-acetyl cysteine protocol and monitoring of her liver enzymes. Other causes of acute liver injury were ruled out. She remained stable during her hospital stay and follow up. Our aim is to highlight the significance of acute liver injury in COVID 19 patients that may lead to fatal outcomes if not managed and monitored accordingly.

LRRC23 truncation impairs radial spoke 3 head assembly and sperm motility underlying male infertility.

Radial spokes (RS) are T-shaped multiprotein complexes on the axonemal microtubules. Repeated RS1, RS2, and RS3 couple the central pair to modulate ciliary and flagellar motility. Despite the cell type specificity of RS3 substructures, their molecular components remain largely unknown. Here, we report that a leucine-rich repeat-containing protein, LRRC23, is an RS3 head component essential for its head assembly and flagellar motility in mammalian spermatozoa. From infertile male patients with defective sperm motility, we identified a splice site variant of LRRC23. A mutant mouse model mimicking this variant produces a truncated LRRC23 at the C-terminus that fails to localize to the sperm tail, causing male infertility due to defective sperm motility. LRRC23 was previously proposed to be an ortholog of the RS stalk protein RSP15. However, we found that purified recombinant LRRC23 interacts with an RS head protein RSPH9, which is abolished by the C-terminal truncation. Evolutionary and structural comparison also shows that LRRC34, not LRRC23, is the RSP15 ortholog. Cryo-electron tomography clearly revealed that the absence of the RS3 head and the sperm-specific RS2-RS3 bridge structure in LRRC23 mutant spermatozoa. Our study provides new insights into the structure and function of RS3 in mammalian spermatozoa and the molecular pathogenicity of LRRC23 underlying reduced sperm motility in infertile human males.

A GHRHR founder mutation causes isolated growth hormone deficiency type IV in a consanguineous Pakistani family.

Background: Isolated growth hormone deficiency (IGHD) is caused by a severe shortage or absence of growth hormone (GH), which results in aberrant growth and development. Patients with IGHD type IV (IGHD4) have a short stature, reduced serum GH levels, and delayed bone age. Objectives: To identify the causative mutation of IGHD in a consanguineous family comprising four affected patients with IGHD4 (MIM#618157) and explore its functional impact in silico. Methods: Clinical and radiological studies were performed to determine the phenotypic spectrum and hormonal profile of the disease, while whole-exome sequencing (WES) and Sanger sequencing were performed to identify the disease-causing mutation. In-silico studies involved protein structural modeling and docking, and molecular dynamic simulation analyses using computational tools. Finally, data from the Qatar Genome Program (QGP) were screened for the presence of the founder variant in the Qatari population. Results: All affected individuals presented with a short stature without gross skeletal anomalies and significantly reduced serum GH levels. Genetic mapping revealed a homozygous nonsense mutation [NM_000823:c.G214T:p.(Glu72*)] in the third exon of the growth-hormone-releasing hormone receptor gene GHRHR (MIM#139191) that was segregated in all patients. The substituted amber codon (UAG) seems to truncate the protein by deleting the C-terminus GPCR domain, thus markedly disturbing the GHRHR receptor and its interaction with the growth hormone-releasing hormone. Conclusion: These data support that a p.Glu72* founder mutation in GHRHR perturbs growth hormone signaling and causes IGHD type IV. In-silico and biochemical analyses support the pathogenic effect of this nonsense mutation, while our comprehensive phenotype and hormonal profiling has established the genotype-phenotype correlation. Based on the current study, early detection of GHRHR may help in better therapeutic intervention.

Laparoscopic Major Vascular Injuries Report of Two Cases and Review.

Laparoscopic surgery is the standard of care for various abdominal pathologies due to its apparent advantages. Unintentional and accidental injuries of the intraabdominal organs and vessels caused by trocars and Veress needles are rare, but potentially fatal complications of laparoscopic surgery. Veress needle and trocar can cause major vascular, bowel, or urinary tract injuries. We report two cases of vascular laparoscopic entry injuries. Laparoscopic entry injuries are rare but life-threatening and can cause multiple organ dysfunctions. Therefore, early recognition, aggressive resuscitation, and appropriate management of laparoscopic entry injuries are vital for better outcomes.

A novel homozygous variant in homologous recombination repair gene ZSWIM7 causes azoospermia in males and primary ovarian insufficiency in females.

Infertility is a common, clinically heterogeneous reproductive disorder worldwide with a prevalence of about 15%. To date about eighty genes have been discovered to cause non-syndromic infertility, affecting males and females equally, though traditionally the genetic analysis of each group has been conducted separately. Here, we report the clinical and genetic characterization of a consanguineous family of Pakistani origin with multiple individuals, including male and female, affected with infertility. Males exhibited non-obstructive azoospermia whereas females had primary ovarian insufficiency. Whole exome sequencing revealed a missense variant [c.176C > T, p. (Ser59Leu)] in the ZSWIM7 gene which functions in homologous recombination repair. The variant was found in a homozygous form in all affected males and females. To our knowledge, this is the first family that has individuals affected with infertility in both sexes. This point to the utility of large consanguineous families with multiple affected siblings to reveal joint mechanisms affecting human reproduction.

A splice site variant in TCTN3 underlies an atypical form of orofaciodigital syndrome IV.

Orofaciodigital syndrome (OFD) is clinically heterogeneous and is characterized by abnormalities in the oral cavity, facial features, digits, and central nervous system. At least 18 subtypes of the condition have been described in the literature. OFD is caused by variants in several genes with overlapping phenotypes. We studied a consanguineous Pakistani family with two affected siblings with an atypical form of OFD type 4 (OFD4). In addition to the typical features of OFD4 that include limb defects and growth retardation, the siblings displayed rare features of scaphocephaly and seizures. Exome sequencing analysis revealed a novel homozygous splice site variant c.257-1G>A in TCTN3 that segregated with disease. This homozygous splice site variant in TCTN3 is most likely the underlying cause of the atypical form of OFD4 observed in this family. Our results contribute to the phenotypic spectrum of TCTN3 associated ciliopathies and will facilitate better clinical diagnosis.

Novel variants in the LRP4 underlying Cenani-Lenz Syndactyly syndrome.

Cenani-Lenz syndrome (CLS) is a rare autosomal-recessive congenital disorder affecting development of distal limbs. It is characterized mainly by syndactyly and/or oligodactyly, renal anomalies, and characteristic facial features. Mutations in the LRP4 gene, located on human chromosome 11p11.2-q13.1, causes the CLS. The gene LRP4 encodes a low-density lipoprotein receptor-related protein-4, which mediates SOST-dependent inhibition of bone formation and Wnt signaling. In the study, presented here, three families of Pakistani origin, segregating CLS in the autosomal recessive manner were clinically and genetically characterized. In two families (A and B), microsatellite-based homozygosity mapping followed by Sanger sequencing identified a novel homozygous missense variant [NM_002334.3: c.295G>C; p.(Asp99His)] in the LRP4 gene. In the third family C, exome sequencing revealed a second novel homozygous missense variant [NM_002334.3: c.1633C>T; p.(Arg545Trp)] in the same gene. To determine the functional relevance of these variants, we tested their ability to inhibit canonical WNT signaling in a luciferase assay. Wild type LRP4 was able to inhibit LRP6-dependent WNT signaling robustly. The two mutants p.(Asp99His) and p.(Arg545Trp) inhibited WNT signaling less effectively, suggesting they reduced LRP4 function.

Eclampsia and posterior reversible encephalopathy syndrome (PRES): A retrospective review of risk factors and outcomes.

Posterior reversible encephalopathy syndrome (PRES) is a clinical and radiological entity initially described in 1996. PRES frequently develops in patients with preeclampsia and eclampsia. There is not much literature on risk factors causing PRES in pregnant patients with eclampsia. This study aimed to determine the incidence of PRES in eclampsia, its association with pregnancy, risk factors, and maternal and perinatal outcomes. PATIENTS AND METHODS: All patients who were admitted with eclampsia and developed PRES in an intensive care unit of a tertiary medical facility between 1997 and 2017 were included in the study. Patients' demographics, pregnancy and gestational data, treatment mode, and outcomes were retrospectively obtained from their medical charts/files. Data were entered using SPSS program version 23. Chi-square test was used to compare the variables, and a p value of < 0.05 was considered statistically significant. RESULTS: A total of 151 patients were admitted during the study period, and 25 developed PRES. The diagnosis was common in patients older than 25 years. Eclampsia patients who developed PRES were without any pregnancy-associated comorbidities (p < 0.08). At the time of diagnosis, their gestational age was more than 36 weeks, which was significant (p < 0.04). Incidence was significantly higher in patients presenting with eclampsia and had recurrent seizures (p < 0.01 and 0.002, respectively). Its incidence was significantly higher in postpartum eclampsia patients (p < 0.01). It was also significantly higher in patients who had cesarean section and hypertension treated with labetalol (p < 0.001 and 0.02, respectively). Overall, the maternal mortality rate of eclampsia patients complicated with PRES was 4% in our population. CONCLUSION: Of eclampsia patients, 16% developed PRES, which is on the lower side than the reviewed literature (10%-90%). Eclampsia on presentation, recurrent seizures, postpartum eclampsia, cesarean delivery, and labetalol use were associated with increased risk of PRES development.

A novel stop-gain mutation in ARMC2 is associated with multiple morphological abnormalities of the sperm flagella.

RESEARCH QUESTION: Male infertility is a global issue worldwide and multiple morphological abnormalities of the sperm flagella (MMAF) is one of the most severe forms of the qualitative sperm defects with a heterogeneous genetic cause that has not been completely understood. Can whole-exome sequencing (WES) reveal novel genetic causes contributing to MMAF in a consanguineous Pakistani family, comprising three infertile brothers? DESIGN: WES and bioinformatic analysis were conducted to screen potential pathogenic variants. The identified variant was validated by Sanger sequencing in all available family members Transmission electron microscopy analyses was carried out to examine the flagella ultrastructure of spermatozoa from patient. RESULTS: WES and Sanger sequencing identified a novel homozygous stop-gain mutation (ENST00000392644.4, c.182C>G, p.S61X) in ARMC2, which is expected to lead to loss of protein functions. Transmission electron microscopy analyses revealed that the flagellar ultrastructure of the patient's spermatozoa was disorganized along with a complete absence of central pair complex (CPC), suggesting that ARMC2 is involved in the assembly, stability of the axonemal complex, or both, particularly the CPC. CONCLUSION: We report that a familial stop-gain mutation in ARMC2 is associated with male infertility in humans caused by MMAF accompanied with loss of CPCs and axonemal disorganization. We provide genetic evidence that ARMC2 is essential for human spermatogenesis and its mutation may be pathogenic for MMAF. These findings will improve the knowledge about the genetic basis of MMAF and provide information for genetic counselling of this disease.