Multiple morphological abnormalities of the flagella (MMAF) represent a severe form of sperm defects leading to asthenozoospermia and male infertility. In this study, we identified a novel homozygous splicing mutation (c.871-4 ACA>A) in the adenylate kinase 7 (AK7) gene by whole-exome sequencing in infertile individuals. Spermatozoa from affected individuals exhibited typical MMAF characteristics, including coiled, bent, short, absent, and irregular flagella. Transmission electron microscopy analysis showed disorganized axonemal structure and abnormal mitochondrial sheets in sperm flagella. Immunofluorescence staining confirmed the absence of AK7 protein from the patients' spermatozoa, validating the pathogenic nature of the mutation. This study provides direct evidence linking the AK7 gene to MMAF-associated asthenozoospermia in humans, expanding the mutational spectrum of AK7 and enhancing our understanding of the genetic basis of male infertility.
Humans ; Male ; Sperm Tail/ultrastructure* ; Homozygote ; Consanguinity ; Asthenozoospermia/pathology* ; Infertility, Male/genetics* ; Mutation ; Pakistan ; Adenylate Kinase/genetics* ; Adult ; Pedigree ; RNA Splicing ; Exome Sequencing ; Spermatozoa

Male infertility can result from impaired sperm motility caused by multiple morphological abnormalities of the flagella (MMAF). Distinct projections encircling the central microtubules of the spermatozoal axoneme play pivotal roles in flagellar bending and spermatozoal movement. Mammalian sperm-associated antigen 17 ( SPAG17 ) encodes a conserved axonemal protein of cilia and flagella, forming part of the C1a projection of the central apparatus, with functions related to ciliary/flagellar motility, skeletal growth, and male fertility. This study investigated two novel homozygous SPAG17 mutations (M1: NM_206996.2, c.829+1G>T, p.Asp212_Glu276del; and M2: c.2120del, p.Leu707*) identified in four infertile patients from two consanguineous Pakistani families. These patients displayed the MMAF phenotype confirmed by Papanicolaou staining and scanning electron microscopy assays of spermatozoa. Quantitative real-time polymerase chain reaction (PCR) of patients' spermatozoa also revealed a significant decrease in SPAG17 mRNA expression, and immunofluorescence staining showed the absence of SPAG17 protein signals along the flagella. However, no apparent ciliary-related symptoms or skeletal malformations were observed in the chest X-rays of any of the patients. Transmission electron microscopy of axoneme cross-sections from the patients showed incomplete C1a projection and a higher frequency of missing microtubule doublets 1 and 9 compared with those from fertile controls. Immunofluorescence staining and Western blot analyses of spermatogenesis-associated protein 17 (SPATA17), a component of the C1a projection, and sperm-associated antigen 6 (SPAG6), a marker of the spring layer, revealed disrupted expression of both proteins in the patients' spermatozoa. Altogether, these findings demonstrated that SPAG17 maintains the integrity of spermatozoal flagellar axoneme, expanding the phenotypic spectrum of SPAG17 mutations in humans.
Humans ; Male ; Infertility, Male/pathology* ; Sperm Tail/ultrastructure* ; Homozygote ; Microtubule-Associated Proteins/genetics* ; Axoneme/genetics* ; Spermatozoa/ultrastructure* ; Adult ; Mutation ; Sperm Motility/genetics* ; Pedigree ; Microtubules ; Microtubule Proteins/genetics*

The syndrome of multiple morphological abnormalities of the sperm flagella (MMAF) is one of the most serious kinds of sperm defects, leading to asthenoteratozoospermia and male infertility. In this study, we use whole-exome sequencing to identify genetic factors that account for male infertility in a patient born from a consanguineous Pakistani couple. A homozygous frameshift mutation (c.1399_1402del; p.Gln468ArgfsTer2) in axonemal dynein light chain domain containing 1 ( AXDND1 ) was identified in the patient. Sanger sequencing data showed that the mutation was cosegregated recessively with male infertility in this family. Papanicolaou staining and scanning electron microscopy analysis of the sperm revealed severely abnormal flagellar morphology in the patient. Immunofluorescence and western blot showed undetectable AXDND1 expression in the sperm of the patient. Transmission electron microscopy analysis showed disorganized sperm axonemal structure in the patient, particularly missing the central pair of microtubules. Immunofluorescence staining showed the absence of sperm-associated antigen 6 (SPAG6) and dynein axonemal light intermediate chain 1 (DNALI1) signals in the sperm flagella of the patient. These findings indicate that AXDND1 is essential for the organization of flagellar axoneme and provide direct evidence that AXDND1 is a MMAF gene in humans, thus expanding the phenotypic spectrum of AXDND1 frameshift mutations.
Humans ; Male ; Sperm Tail/ultrastructure* ; Frameshift Mutation ; Infertility, Male/pathology* ; Pakistan ; Pedigree ; Consanguinity ; Axonemal Dyneins/genetics* ; Adult ; Spermatozoa ; Exome Sequencing

Objective: To evaluate the antioxidant potential and pancreatic lipase inhibitory action of optimized hydroethanolic extracts of Solanum nigrum. Methods: Optimized extraction for maximum recovery of metabolites was performed using a combination of freeze-drying and ultrasonication followed by determination of antioxidant and antiobesity properties. The ultra-high performance liquid chromatography equipped with mass spectrometry was used to analyze metabolite profiling of Solanum nigrum. Computational studies were performed using molecular docking and electrostatic potential analysis for individual compounds. The hypolipidemic potential of the most potent extract was assessed in the obese mice fed on fat rich diet. Results: The 80% hydroethanolic extract exhibited the highest extract yield, total phenolic contents, total flavonoid contents along with the strongest 2,2-diphenyl-1-picrylhydrazyl scavenging activity, total antioxidant power, and pancreatic lipase inhibitory properties. The 80% hydroethanolic extract not only regulated the lipid profile of obese mice but also restricted the weight gain in the liver, kidney, and heart. The 80% hydroethanolic extract also reduced alanine transaminase and aspartate transaminase concentrations in serum. The effects of plant extract at 300 mg/kg body weight were quite comparable with the standard drug orlistat. Conclusions: Solanum nigrum is proved as an excellent and potent source of secondary metabolites that might be responsible for obesity mitigation.

Objective: To profile the secondary metabolites and to evaluate the antidiabetic potential of hydroethanolic leaf extracts of Conocarpus lancifolius. Methods: The various hydroethanolic extracts of Conocarpus lancifolius leaf were prepared by ultrasonication assisted freeze-drying. Total phenolic contents, flavonoid contents, antioxidant activity, α-glucosidase and α-amylase inhibitions of leaf extracts were determined. The metabolite profiling was accomplished by UHPLC-Q-TOF-MS/MS analysis. The antidiabetic assessment of the most potent extract was carried out by measuring the hypoglycemic and hypolipidemic effect in the high fat diet-fed diabetic albino mice. The blood glucose level, haemoglobin, total cholesterol, high-density lipoproteins (HDL) and low-density lipoproteins (LDL) were determined. Results: The 60% ethanolic extract exhibited the highest phenolic and flavonoid contents of (349.39 ± 2.13) mg GAE/g dry extract and (116.95 ± 2.34) mg RE/g dry extracts, respectively, and the highest DPPH scavenging activity with an IC50 value of (32.87 ± 1.11) μg/mL. The IC50 values for α-glucosidase and α-amylase inhibitions were (38.64 ± 0.93) μg/mL and (44.80 ± 1.57) μg/mL, respectively. UHPLC-Q-TOF-MS/MS analysis confirmed the presence of gallic acid, ellagic acid, corilagin, kaempherol-3-O-rutinoside, caffeic acid derivative, isorhamnetin and galloyl derivatives in the 60% ethanolic extract. Plant extract at a dose of 450 mg/kg body weight reduced blood glucose level, total cholesterol, LDL and HDL, and increased haemoglobin in alloxan-induced diabetic mice, Conclusions: Conocarpus lancifolius leaves are proved as a good source of biologically functional metabolites and possess antidiabetic activity which may be further explored to treat diabetes.

Objective: To evaluate the potential of both jamun (Syzygium cumini) seed and fruit extracts against hyperglycemia. Methods: Male Sprague Dawley rats were used to evaluate hypoglycemic potential of jamun extracts. Purposely, jamun fruit and seed's ethanolic extracts based diets were provided to normal and high sucrose diet induced hyperglycemic/diabetic rats for sixty days. The serum glucose and insulin levels were monitored at monthly intervals to evaluate hypoglycemic effect of jamun extracts. Results: The results of instant research depicted that both seed and fruit extracts reduce the blood glucose level significantly and also regulate the insulin levels in hyperglycemic rats. It was noted that jamun fruit extract attenuated serum glucose levels to 5.35％and 12.29％ in normal and hyperglycemic rats, respectively; while insulin levels were improved by 2.82％ and 6.19％, correspondingly. Whereas, jamun seed extract reduced glucose to 7.04％&14.36％and showed 3.56％&7.24％higher insulin levels in normal&hyperglycemic rats, respectively. Conclusions: The present research revealed that both jamun fruit and seeds have potent prophylactic role against hyperglycemia. In this respect, diet based regimen may be tailored using jamun fruit/seed and their extracts to alleviate hyperglycemia.

Objective To evaluate the potential of both jamun (Syzygium cumini) seed and fruit extracts against hyperglycemia. Methods Male Sprague Dawley rats were used to evaluate hypoglycemic potential of jamun extracts. Purposely, jamun fruit and seed's ethanolic extracts based diets were provided to normal and high sucrose diet induced hyperglycemic/diabetic rats for sixty days. The serum glucose and insulin levels were monitored at monthly intervals to evaluate hypoglycemic effect of jamun extracts. Results The results of instant research depicted that both seed and fruit extracts reduce the blood glucose level significantly and also regulate the insulin levels in hyperglycemic rats. It was noted that jamun fruit extract attenuated serum glucose levels to 5.35% and 12.29% in normal and hyperglycemic rats, respectively; while insulin levels were improved by 2.82% and 6.19%, correspondingly. Whereas, jamun seed extract reduced glucose to 7.04% & 14.36% and showed 3.56% & 7.24% higher insulin levels in normal & hyperglycemic rats, respectively. Conclusions The present research revealed that both jamun fruit and seeds have potent prophylactic role against hyperglycemia. In this respect, diet based regimen may be tailored using jamun fruit/seed and their extracts to alleviate hyperglycemia.

Poultry farming not only provides high nutritious food but also creates employment opportunity for rural masses. Documented evidences elaborates that helminth parasitism is most deciduous problem of chickens especially in developing world. Ascaridia (A.) galli, a nematode of small intestine, has been considered as the most common and important parasite of chicken. The present study was carried out to investigate prevalence and severity of A. galli in White Leghorn layers (housing type: battery cage and deep litter, 50 each) and Fayoumi-Rhode Island Red crossbred (male and female: 50 each) flock rearing at Government Poultry Farm, Dina, Punjab, Pakistan. Two hundred faecal samples were examined by using standard parasitological and McMaster egg counting technique. The overall prevalence was 24.5% at farm, 13% in White leghorn layer (battery cage=2%, deep litter=24%) and 36% in Fayoumi-Rhode Island Red (male=34%, female=38%). It was also observed that White leghorn layer rearing in deep litter had more severe infection (EPG=1920) of A. galli compare with battery cages birds (EPG=500). Parasite prevalence was significantly related with sex (P<0.05) in Fayoumi-Rhode Island Red and male birds had less number of average parasites (0.34±0.47) as compared to females (0.38±0.490). Additionally, female birds were under serious threat of infection (EPG=2270) compared with its counterpart (EPG=1250). Given the high infection rates, particular attention should be paid to management and provision of feed supplement to White leghorn layer housing in deep litter and female bird of Fayoumi-Rhode Island Red crossbred.