Factor Gene Research Articles

Detection of single-nucleotide polymorphisms in growth hormone and insulin-like growth factor-1 genes related to growth traits in purebred and crossbred quails.

There is a limited amount of research conducted on quail breeding domestically and internationally, particularly at the molecular level. This study aimed to detect single-nucleotide polymorphisms in the growth hormone (GH) and insulin-like growth factor-1 (IGF-1) genes across two quail varieties and their hybrids correlate these genetic factors with body weight (BW) and growth rate at 0 and 6 weeks, and assess crossing effects. White and Japanese quail were crossed. Simultaneously producing pure varieties and crosses (genotypes) was achieved through this breeding strategy. Fifty females from each genotype were randomly selected for blood sampling. Genomic DNA was extracted and amplified from the blood using the DNeasy blood kit (Qiagen, Germany). Nucleotide polymorphism between quail genotypes was determined through DNA sequencing. Two types of alleles (A and B) for the GH gene in quails showed significant genotypic differences (AA, BB, and AB). The quail carried a mutated IGF-1 gene. For growth traits, substantial positive heterosis was detected. The genotype AA had the highest BW and weight gain. The white variety can act as a sire, and both white and Japanese varieties can function as dams to improve growth traits. The growth characteristics of the hybrids surpassed those of the original varieties.

First report of Nigrospora chinensis causing dragon fruit (Selenicereus monacanthus) stem spot in China.

Dragon fruit (Selenicereus monacanthus), renowned for its economic value and dual utility in both culinary and medicinal applications, is predominantly cultivated in China. In July 2023, a stem spot disease was found on dragon fruit ("Zi honglong" cultivar) plants with 37% incidence, in Huajiang Town (N25°40', E105°39'), Guanling County, Anshun City, Guizhou Province. The symptoms appeared as yellow spots surrounded by watery stains, then the spots expanded to suborbicular, which finally led stem to wither. Twelve symptomatic stem samples were collected in a 1.3-hectare plantation and cut into small pieces (5 mm × 5 mm), sterilized the surface with 75% ethanol for 30 seconds, washed 3 times with disinfected distilled water, moved to potato dextrose agar (PDA) medium, and incubated at 28°C for 5 days. Once the mycelium had developed, they were transferred to another PDA medium and cultured at 28°C for a period of 3-5 days. Totally fifteen identical strains were isolated, their colonies were white and round in shape; hyphae were smooth, hyaline; conidia were globose or subglobose, smooth, aseptate, 5.8-11.9 × 4.2-10.6 μm (av. = 8.8 × 7.4 μm, n = 30), light brown in early stage and gradually turning black over time; sterile cells were terminal on hyphae, pale to dark brown, uhceiform or oval, 17.5-21.6 × 4.6-8.7 μm (av. = 19.4 × 5.8 μm, n = 30). The morphologic characteristics of the isolates matched Nigrospora chinensis described by Wang etal.(2017). The PCR amplification was carried out by 3 primers ITS1/ITS4 (Vilgalys et al. 1990; White et al. 1990), EF1-728/EF2 (O'Donnell et al. 1998; Carbone et al. 1999) and BT2A/BT2B (Glass et al. 1995) belonged to the internal transcribed spacer (ITS), translation elongation factor-1 (TEF1) and β-tubulin (TUB2) gene loci, respectively. The sequences of a representative strain (GUCC 524) had 99.38% (ITS: 483/486 bp, PP391347 vs KX985970), 99.79% (TEF1: 482/483 bp, PP400678 vs KY019427) and 99.73% (TUB2: 372/373 bp, PP400677 vs KY019497) identities with those of N. chinensis (strain LC 3085). The phylogenetic tree constructed by three gene combinations showed that GUCC 524 was significantly clustered with N. chinensis. Ten 6-month-old dragon fruit ("Zi honglong" cultivar) seedlings were engrafted with 10 μL conidial suspension (1×105 conidia/mL), packaged with sealed film, two of them were inoculated with sterile distilled water as controls, and placed in a greenhouse at 28℃ for 10 days, inoculated plants showed yellow spots analogous to field symptoms, no symptoms were found in control plants. This experiment was repeated three times. Morphological character and molecular identification based on 3 gene loci of the strains isolated from the inoculated stems, were consistent with those of the original isolated strains. Therefore, based on morphological identification, phylogenetic analysis and pathogenicity test, the pathogen was identified as N. chinensis. Our study firstly reported N. chinensis as a pathogen causing stem spot disease on dragon fruit. N. chinensis is an important agent resulting for economic losses, previously reported on Camellia sinensis (Wang et al. 2017), Saccharum officinarum (Raza et al. 2019), Aucuba japonica (Qin et al. 2021). This report establishes a pivotal reference point for the progression of scientific strategies in preventing and controlling this disease associated with N. chinensis.

Nutritional synergy: optimizing growth in Siruvidai chicken via diverse dietary energy and protein strategies and gene expression analysis - brooder phase.

Nutrient supply regulates overall body growth directly or indirectly through its influence on regulatory factors optimizing nutrient requirements becomes crucial before embarking on genetic improvements. Hence this study addresses this gap by evaluating the effect of feeding varying energy and crude protein levels on growth performance and gene expression related to the growth of indigenous Siruvidai chicken from 0 to 12 weeks. A 360-day-old straight-run Siruvidai chick were randomly distributed into six experimental groups with three replicates of each 20 chicks. The birds were fed corn-soy-based diets formulated with two levels of energy (2500 and 2700kcal ME/kg) each with three levels of crude protein (16, 18, and 20%) during the brooder stage (0-12 weeks) in 2 × 3 factorial design. Results revealed that there was no significant effect on the energy and protein interaction levels on average feed intake, body weight gain and feed conversion ratio in Siruvidai chicken at 12 weeks. The results showed significantly (P < 0.05) lower feed intake in 18% protein fed groups and significantly (P < 0.01) lower feed intake in higher energy 2700kcal ME/kg fed groups. A better feed conversion ratio (4.06 and 4.21) was observed on the effect of protein levels in bird diets with 18% and 20% protein fed groups. The Growth Hormone (GH) and Myostatin (MSTN) gene expression were significantly (P < 0.01) higher in 16% CP and 2500kcal ME/kg in hepatic tissue. The high protein and low energy diet up-regulated the Insulin-like Growth Factor-1 (IGF-1) gene expression in hepatic tissue. The study concluded that Siruvidai chicken fed with 18% crude protein and 2500kcal ME/kg is optimum for 0-12 weeks of age.

Dietary chromium-methionine supplementation and broiler (22–43 days) responses during heat stress. 2 - Physiological variables, and heat shock protein 70 and insulin-like growth factor-1 gene expression

Dietary chromium-methionine supplementation and broiler (22–43 days) responses during heat stress. 2 - Physiological variables, and heat shock protein 70 and insulin-like growth factor-1 gene expression

Characterization of missense nonsynonymous single-nucleotide polymorphism of runt-related transcription factor-2 gene - An in silico approach.

Single-nucleotide polymorphism (SNP) codes for multiple amino acids, impacting protein functions and disease prognosis. Runt-related transcription factor-2 (RUNX2), a transcription factor linked to osteoblast differentiation, regulates cell proliferation in endothelium and osteoblastic cells. Understanding Runx2's role in nonosseous tissues is rapidly advancing. This study aims to identify harmful SNPs of the RUNX2 gene that may alter disease susceptibility using computational techniques. The study uses various in silico methods to identify nonsynonymous SNPs (nsSNPs) of the RUNX2 gene, which could potentially alter protein structure and functions, with further analyses by I-Mutant, ConSurf, Netsurf 3.0, GeneMANIA, and Have (y)Our Protein Explained. Six missense nsSNPs were identified as potentially harmful, disease-causing, and damaging. Four were found to be unstable, while five were conserved. All six nsSNPs had a coiled secondary structure. Five nsSNPs were found to be destabilized. The RUNX2 gene's deleterious missense nsSNPs were identified by this study, and they may be exploited in future experimental studies. These high-risk nsSNPs might be considered target molecules in therapeutic and diagnostic therapies in teeth and bone development.

Multi-omics revealed the mechanism of feed efficiency in sheep by the combined action of the host and rumen microbiota

This study was conducted to investigate potential regulatory mechanisms of feed efficiency (FE) in sheep by linking rumen microbiota with its host by the multi-omics analysis. One hundred and ninety-eight hybrid female sheep (initial body weight = 30.88 ± 4.57 kg; 4-month-old) were selected as candidate sheep. Each test sheep was fed in an individual pen for 60 days, and the residual feed intake (RFI) was calculated. The ten candidate sheep with the highest RFI were divided into the Low-FE group, and the ten with the lowest RFI were divided into the High-FE group, all selected for sample collection. The RFI, average daily gain and average daily feed intake were highly significantly different between the two experimental groups (P < 0.05). Compared with Low-FE group, the insulin-like growth factor-1 and very low-density lipoprotein in serum and the propionate in rumen significantly increased in High-FE group (P < 0.01), but the acetate:propionate ratio in rumen significantly decreased in High-FE group (P = 0.034). Metagenomics revealed Selenomonas ruminantium, Selenomonas sp. and Faecalibacterium prausnitzii were key bacteria, and increased abundance of the genes encoding the enzymes for cellulose degradation and production of propionate in High-FE group. The results of proteomics and section showed the rumen papilla length (P < 0.001) and expression of carbonic anhydrase and Na+/K+-ATPase were significantly higher in High-FE group (P < 0.05). On the other hand, the acetyl-CoA content significantly increased in the liver of High-FE group (P = 0.002). The relative expression levels of insulin-like growth factor-1 and apolipoprotein A4 genes were significantly up-regulated in the liver of High-FE group (P < 0.01), but relative expression level of monoacylglycerol O-acyltransferase 3 gene was significantly down-regulated (P = 0.037). These findings provide the mechanism by which the collaborative interaction between rumen microbiota fermentation and host uptake and metabolism of fermentation products impacts feed efficiency traits in sheep.

Effect of dietary chitosan on the growth performance, intestinal histology and growth-related gene expression in stellate sturgeon (Acipenser stellatus) juveniles.

As sturgeon breeding has proliferated, there has been a heightened demand for growth stimulators in their diets. This study aimed to determine the impact of dietary chitosan on growth performance, whole-body proximate composition, growth-related gene expression, and intestinal histology in juvenile Acipenser stellatus. A total of 180 A. stellatus juveniles with an average weight of 31.90 ± 0.73 g were fed with diets containing 0 (control), 1.5, 3.0, 4.5, and 6.0 g chitosan.kg-1 basic diet for eight weeks. The findings revealed a significant enhancement in growth performance with rising chitosan concentrations. Furthermore, chitosan supplementation upregulated the expression of the growth hormone gene in both brain and liver tissues. In liver samples, the most pronounced expression of the insulin-like growth factor-1 gene was noted at 6.0 g chitosan.kg-1, while in brain samples, peak expressions were observed in both the 4.5 and 6.0 g chitosan.kg-1 treatments. While the whole-body proximate composition remained relatively stable, there was a notable decrease in whole-body lipids with the escalation of chitosan dosage. Intestinal villi dimensions, both height and width, were amplified in the chitosan-supplemented groups compared to controls. In summation, chitosan supplementation showed promise in bolstering growth performance, refining intestinal morphology, and enhancing growth-related gene expression. Analysis of the polynomial regression of weight gain and specific growth rate revealed that the optimum dietary chitosan requirements in A. stellatus were 5.32 and 5.21 g chitosan.kg-1, respectively.

G-quadruplex is involved in the regulation of BmSGF1 expression in the Silkworm, Bombyx mori.

Advanced DNA structures, such as the G-quadruplex (G4) and the i-motif, are widely but not randomly present in the genomes of many organisms. A G4 structure was identified in the promoter of the silk gland factor-1 gene (SGF1), which is the main regulatory gene for silk production in Bombyx mori. In this study, a BmSGF1 G4-/- homozygous mutant was generated with the G4 sequence knocked out. The promoter activity of BmSGF1 was lowered in the BmSGF1G4-/- mutant. Pyridostatin (PDS) stabilized the G4 structure and increased the promoter activity of BmSGF1, whereas anti-sense oligonucleotide (ASO) complementary to the G4 sequence suppressed the promoter activity of BmSGF1. Compared with wild-type larvae, the deletion of the BmSGF1 G4 structure decreased both the expression of BmSGF1 and the fibroin heavy chain gene BmFib-H in the posterior silk gland and the weight of the cocoons. Overall, these results suggest that the promoter G4 structure of BmSGF1 participates in the transcription regulation of the BmSGF1 gene in the silkworm.

A High-Quality Blue Whale Genome, Segmental Duplications, and Historical Demography.

The blue whale, Balaenoptera musculus, is the largest animal known to have ever existed, making it an important case study in longevity and resistance to cancer. To further this and other blue whale-related research, we report a reference-quality, long-read-based genome assembly of this fascinating species. We assembled the genome from PacBio long reads and utilized Illumina/10×, optical maps, and Hi-C data for scaffolding, polishing, and manual curation. We also provided long read RNA-seq data to facilitate the annotation of the assembly by NCBI and Ensembl. Additionally, we annotated both haplotypes using TOGA and measured the genome size by flow cytometry. We then compared the blue whale genome with other cetaceans and artiodactyls, including vaquita (Phocoena sinus), the world's smallest cetacean, to investigate blue whale's unique biological traits. We found a dramatic amplification of several genes in the blue whale genome resulting from a recent burst in segmental duplications, though the possible connection between this amplification and giant body size requires further study. We also discovered sites in the insulin-like growth factor-1 gene correlated with body size in cetaceans. Finally, using our assembly to examine the heterozygosity and historical demography of Pacific and Atlantic blue whale populations, we found that the genomes of both populations are highly heterozygous and that their genetic isolation dates to the last interglacial period. Taken together, these results indicate how a high-quality, annotated blue whale genome will serve as an important resource for biology, evolution, and conservation research.

TCM formula for trauma treatment screening and its role of promoting infectious wound coalescence investigating

In pet clinics, the number of cases using trauma drugs accounts for >10% of the total number of cases, and most wounds are healing by second intention. The prolongation of wound healing time causes inconvenience and burden to pets and pet owners. Therefore, how to reduce wound healing time and achieve maximum recovery of tissue function and aesthetics is one of the focuses of veterinary clinical practice. Wound suppuration caused by Staphylococcus aureus and Pseudomonas aeruginosa is the main cause of delaying wound healing. Clinically, available antimicrobial treatments are almost exhausted due to the production of large numbers of resistant bacteria. At present, there are no bacteria resistant to traditional Chinese medicine (TCM), which makes TCM have the potential to become an effective drug for the treatment of bacterial infections, so the use of TCM in the treatment of traumatic infections has broad prospects. Based on the characteristics of infection syndrome, three different prescriptions were formulated in our laboratory, and the most effective prescription and dosage form was screened and named Lianrong Healing Cream (LRHC). The results showed that LRHC regulated the expression of fibroblast growth factor-2 (FGF-2), epidermal growth factor-1 (EGF-1), transforming growth factor-β (TGF-β) and vascular endothelial growth factor-1 (VEGF-1) genes in wound tissues and fibroblasts, thereby accelerating wound healing and repairing wound appearance and function. The results of this study may be help to develop TCM formulation for traumatic infections.

Identification of genetic variants of bGH, GHR and IGF1 gene and their association with lactation persistency in Tharparkar cattle

Persistency can be explained as number of days during which the high level of milk production is maintained after the peak milk yield. The aim of this study was to identify genetic variants of bovine-Growth Hormone (bGH), Growth Hormone Receptor (GHR) and Insulin like Growth Factor1 (IGF1) gene and their association with lactation persistency in Tharparkar cattle. Study was carried out using data of daily milk yield, monthly test days, total milk yield and peak yield of 372 lactation records of 190 Tharparkar cattle maintained at Livestock Research Centre of ICAR-National Dairy Research Institute from 1996-2019. Lactation persistency was estimated using Woods and Wilminks method and its least squares means were 2.37±0.08 and -5.4±0.56, respectively. Investigations on polymorphism were performed using seven reported primers for amplifying targeted regions of bGH, GHR and IGF1 gene. In PCR-RFLP, bGH-MspI RFLP had shown polymorphism in intron 3 and its genotypic frequencies were TT 0.49, CT 0.39 and CC 0.12. The allelic frequency of C and T were 0.31 and 0.69, respectively. The bGH-AluI RFLP had shown monomorphism in exon 5. GHR-AluI had shown polymorphism in exon 10 with only two genotypes AA and AG with frequency of 0.92 and 0.08, respectively. The frequency of A and G allele was 0.96 and 0.04, respectively. In GHR-StuI and IGF1-TasI in promoter region, RsaI in exon 3 and SnaB I in exon 1 had shown monomorphism. No significant association was found between genetic variants of bGH and GHR with lactation persistency in Tharparkar cattle.

First report of leaf spot disease caused by Alternaria alternata on Polygonatum cyrtonema Hua in Hunan Province of China.

Polygonatum cyrtonema Hua, a perennial plant of the Asparagaceae family, is an important herb in Chinese medicine and is mainly grown in the Chinese provinces of Guizhou, Hunan, Yunnan, Anhui, and Zhejiang (Chen et al. 2021). In June 2021, a new case of leaf spot disease was detected in an 80 m2 plantation of P. cyrtonema on Xuefeng Mountain, Huaihua City, Hunan Province (27°17'30″N, 110°24'20″E). It infected almost 40% of the total planted area. Initially, irregular light brown spots appeared on the leaves, gradually turning dark brown and coalescing to form large necrotic areas, after which the affected plant turned yellow and eventually died. Ten disease samples were collected from ten plants in the plantation area. The leading edge of necrotic tissues were rinsed with sterile water and then disinfected with 3% hydrogen peroxide for 30 s, followed by 75% ethanol for 90 s, and rinsed three times with sterile water. Samples were then placed on water agar plates and incubated in the dark in a constant temperature incubator at 28 ℃ for 3-5 days. After mycelial growth was observed in the media, the hyphae were transferred to potato dextrose agar plates and incubated for 3-5 days at 28 ℃ in the dark. Ultimately, 12 purified fungal isolates were obtained, some of which were morphologically similar, including 10 that were Alternaria (83.3% isolation rate). Three representative isolates (HJYB1, HJYB2, and HJYB3) were selected for further study. The initial colonies were grayish green with white fluffy mycelia on the surface and a prominent white rim, which became brown with dense, cottony aerial mycelia as the colonies matured. The conidia were obpyriform or ellipsoidal, pale to dark brown, with 0-4 transverse and 0-3 longitudinal septa, some with a short cylindrical beak at the tip. They measured 11.826-28.873 × 6.231-26.018 μm (n = 100). To further confirm the identity of the isolates, their rDNA internal transcribed spacer region (ITS), β-microtubulin (TUB2) and translation elongation factor-1 (TEF-1) genes were amplified and sequenced using the ITS4/ITS5, TUB2F/R and EF-526F/1567R primers, respectively (Hong et al. 2006). The sequences were submitted to GenBank (ITS: OR513924, OR513964, OR519874; TUB2: OR526928, OR533421, OR526929; TEF: OR526926, OR533420, OR526927). A concatenated phylogenetic tree of the three genes showed that the isolate clustered significantly with Alternaria alternata. Based on morphological identification and phylogenetic tree analysis, the isolate was identified as A. alternata. We carried out pathogenicity tests on four uniformly growing P. cyrtonema plants. Three of these plants were used as experimental plants and one as a control. For each plant, three young leaves were selected and inoculated with 6 × 6 mm PDA blocks, while sterile PDA blocks were used as controls. The treated plants were subjected to 10 days of stable temperature in a climatic chamber set at 28°C, 80% constant relative humidity and 12 hours of light per day. The pathogenic lesions appeared and the pathogens re-isolated from the diseased leaves showed similar morphological characteristics to representative isolates and were confirmed as A. alternata by DNA sequencing, thus fulfilling Koch's postulates. A. alternata is the major causal agent of leaf spot on P. sibiricum (Zou et al. 2023) and Agrimonia pilosa (Jiang et al. 2023). As far as we know, leaf necrosis caused by A. tenuissima has been found on P. cyrtonema (Li et al. 2020). To our knowledge, this is the first report of A. alternata causing leaf spot disease in P. cyrtonema. These findings form the basis for the management of this leaf spot disease.

Dietary Effect of S-Methylmethionine Sulfonium Chloride on Growth, Serum Biochemical Parameters, Body Composition, and Expression of Some Related Genes in Oreochromis niloticus

Abstract The main objective of the current trial was to investigate the impacts of tilapia diets supplemented with S-methylmethionine sulfonium chloride (MMSC) on the growth performance, feed efficiency, body analysis, blood biochemistry and regulation of myostatin (MSTN) and insulin-like growth factor-1 (IGF-1) genes of nile tilapia (Oreochromis niloticus). The experimental fish (n=180) weighing 22.4±0.17 g were allocated randomly into three equal groups in triplicate as follows; the control group received an un-supplemented diet; the other two groups received two different levels of MMSC (0.2 and 0.4 g/kg) for eight weeks. The obtained findings demonstrated that tilapia diets enriched with small dosages of MMSC considerably (P&lt;0.05) enhanced all assessed growth performance and feed efficiency indicators when compared with the control group. When compared to the control group, tilapia meals supplemented with 0.2 g MMSC significantly (P&lt;0.05 or 0.01) raised blood protein profile, particularly total protein and globulin levels. Contrarily, blood creatinine levels were significantly (P&lt;0.01) reduced in the group of tilapia fed diets containing MMSC (0.2 or 0.4 g/kg) compared to the group without receiving any supplementation. Whereas, fish body analysis showed higher significant increases in crude protein and ether extract levels (P&lt;0.01) as well as reduced ash content (P&lt;0.05) in the fish group that received lower MMSC doses in comparison to other treated and control groups. In addition, MMSC dietary supplementation significantly downregulated the expression of MSTN and upregulated IGF-1 mRNA expression compared with the control group. Additionally, both dosages of MMSC supplementation modestly enhanced the intestinal villus histomorphometric score indices with observed tall, thick, and various broad tips in comparison to the control group. In conclusion, it could be recommended that fish diets supplemented with 0.2 g MMSC per kg diet may promote the growth and general health status of Nile Tilapia.

Polymorphism analysis of the Insulin-like Growth Factor-1 (IGF-1) gene encoding growth in F5 Golden Kamper chicken (Gallus gallus domesticus)

Golden Kamper Chicken is a chicken cross between pelung and layer chickens with good meat and egg productivity. Insulin-like growth factor-1 (IGF-1) gene is one factor affecting the speed of chicken growth. This research aims to study the character of F5 Golden Kamper chickens and analyze the IGF-1 gene to determine its effect on chicken growth. The research will be done by hybridizing the F4 Golden Kamper hen and rooster, raising brood stock and DOC, egg collection, growth data collection, blood collection, DNA isolation, DNA amplification, electrophoresis, and sequencing. The observed parameters are DOC weight for the first seven weeks and polymorphism of the IGF-1 gene. Data was analyzed using Microsoft Excel, Genestudio, MEGA11, and IBM SPSS 25 software. Sequencing data were processed using the Genestudio application, alignment with MEGA11, and genotype-haplotype associations using Pearson correlation and Linear Regression F test. The results showed that the molecular marker polymorphism of the IGF-1 gene was found at 12 mutation points. IGF-1 gene polymorphism did not correlate with the weight of F5 Golden Kamper chickens.

The polymorphism of the ovine insulin like growth factor-2 (IGF2) gene and their associations with growth related traits in Tibetan sheep.

In the current study, the role of the ovine IGF2 as a potential candidate gene was investigated as though marker-assisted selection in Chinese Tibetan sheep. The Sanger DNA sequencing method explored five single nucleotide polymorphisms (SNPs) in 5'UTR of the ovine IGF2 gene (C15640T, G15801A, G15870A, C15982G and G15991A) in Chinese Tibetan sheep. The frequencies of four SNPs were within the Hardy-Weinberg Equilibrium (chi-square test) except C15982G. The statistical analysis indicated that the C15640T and G15801A were significantly associated with body height, body length, chest circumference, and body weight (P < 0.05 or P < 0.01). Furthermore, C15982G variant exhibited significant correlation with the body weight (P < 0.01). These findings suggests that the promoter variants of IGF2 gene could be used as a candidate gene through marker-assisted selection for the body weight and body measurement traits in Tibetan sheep breeding program.

Intraarticular Delivery of AAV5-Mir-125a-5p Regulates Key Molecular Mediators of Hemophilic Arthropathy

Background: Hemophilic arthropathy (HA) is a major comorbidity in patients with hemophilia characterized by chronic inflammation of the joints, stiffness and loss of function. The expression of microRNAs are known to be altered in various joint diseases such as rheumatoid arthritis and osteoarthritis which share clinical attributes with HA. One such microRNA is miR-125a-5p, known to be involved in regulating inflammatory mediators in the arthritic joints. The current study was designed to investigate the role of miR125a-5p in HA and examine the phenotypic effect of its targeted reconstitution in the joints. Methods: We utilized a chronic arthropathy model in hemophilia A mice to study the impact of miR125a-5p in disease progression. First, we performed a targeted screen of miR125a-5p and its target genes (STAT1 and TRAF6) using digital PCR (dPCR). Further, to validate if reconstitution of miR125a-5p aids in modulating the molecular mediators of HA, we performed the Adeno-associated virus vector (AAV) serotype 5 targeted delivery of miR125a (AAV5-miR125a) either alone (2.13x10 11vgs/joint) or in combination with capsid-modified AAV8-human Factor 8 (F8)(AAV8-F8) gene therapy at a low dose (1x10 11vgs/animal) intravenously in F8 tm1kaz/Jmice. Briefly, the animals were administered with vectors (alone or in combination), and seven days post vector administration, the animals were subjected to multiple bleeding episodes in the right knee (Day 0, 14, and 30) to mimic chronic hemarthrosis. On day 45, blood samples were collected from experimental animals for FVIII specific clotting assay, and joint tissues were isolated to perform further molecular and histochemical analysis. Results: Our dPCR analysis revealed a significant reduction (2-fold) of miR125a-5p (3518 copies/µl vs 7242 copies/µl, p≤0.0001) in injured joints compared to control joints. Absolute quantification of target genes, namely STAT1 and TRAF6 showed significantly higher expression in injured joints (7.6 to 10.6 fold, respectively) when compared to control joints. Further immunohistochemical examination of joint sections from the experimental mice confirmed the elevated expression of these immunological mediators (STAT1, TRAF6). Upon administration of AAV vectors containing miR125a and/or F8, we observed a significantly increased expression of miR125a-5p (~12 fold) in only miR125a vector-administered animals while the combination vector treated group had, ~44-fold higher levels of miR125a-5p. The coagulant assay revealed an increased FVIII activity (~17%) in only F8 vector-treated animals and approximately 20% FVIII levels in mice that received both miR125a and F8 vectors at low doses. Surprisingly, the animals that received only F8 vectors also showed significantly upregulated miR125a-5p expression (~20 fold) signifying the role of miR125a-5p in HA pathology. In line with the above findings, immunohistochemical analysis of joint tissues revealed decreased expression of immunological markers STAT1, TRAF6 (direct targets of miR125a-5p), and the chondro-degenerative matrix metalloproteinases 3, 9, 13 in vector-treated groups when compared to injured joints with no treatment. Hematoxylin and Eosin (H&amp;E) staining of joint tissues from experimental animals was performed which revealed that animals that received both miR125a and F8 vectors showed decreased synovial hyperplasia, villi formation, cartilage erosion, and reduced hemosiderin deposition signifying moderate phenotype rescue. Conclusions: Our data confirms that miR125a-5p is a major molecular target that regulates inflammatory response in chronic HA. Targeted delivery of miR125a in tandem with F8 gene augmentation had an impact on several molecular mediators like STAT1, TRAF6, MMP3,9, and 13. The reconstitution of miR125a and a higher dose of F8 vectors might be beneficial to reverse chronic HA.

Deciphering the Role of Insulin-Like Growth Factor 1 in Endometrial Cancer in Patients With Polycystic Ovary Syndrome: Protocol for a Methodological Approach Using Cell Culture Experiments.

Endometrial cancer (EC) is the most common gynecological cancer in women globally. It is linked to increasing obesity rates and longer life spans. The molecular mechanisms leading to EC are unclear; however, women with polycystic ovary syndrome (PCOS) have a 3- to 5-fold increased EC risk. According to a pilot study conducted in the United Kingdom, insulin-like growth factor-1 (IGF-1) gene and protein were raised in the endometrium and blood of women with EC and PCOS, compared with those without PCOS (controls). Therefore, raised serum IGF-1 levels may contribute to an increased EC risk in women with PCOS, but it is necessary to test this hypothesis since not all studies have demonstrated this association. This study aims to investigate the role of IGF-1 in mediating EC risk in PCOS. This will be achieved by evaluating the proliferative effects of PCOS serum, IGF-1, and IGF-1 antagonist on human endometrial cancer 1-A and 1-B cell lines, with a comparison to controls (using serum from women without PCOS and cell culture media). The study will also identify differentially expressed genes and pathways activated by various treatments. We intend to recruit 20 women with PCOS and 20 women without PCOS for this cross-sectional study. All experiments will be carried out 4 times to ensure consistency. We will perform transcriptomic and phosphoproteomic profiling to identify differentially expressed genes and phosphoproteins between different treatments using RNA sequencing and phosphoproteomics. We will also perform bioinformatics pathway analysis to identify whether any unique collection of genes or phosphoproteins explains increased EC risk in PCOS. The primary outcome measure will be the cell proliferation (growth) difference measured by cell index values. Our protocol stands out due to its unique approach; no previous study has used this approach to investigate the oncogenic effect of serum from women with PCOS. Additionally, no previous study has considered the differential mutations of genes related to the insulin signaling pathway across various types of human EC cell lines and the potential impact of these variations on their experimental findings. Participants are currently being recruited. It is expected that preliminary findings suitable for analysis and publication will be available by the summer of 2024. Although we currently do not have any results to report, sharing our protocol at this stage will aid in research collaboration, provide an opportunity for early feedback, and help reduce duplication of effort by other research groups. The findings of our study will have broader implications. A deeper understanding of the mechanisms underlying the regulation of the IGF system in PCOS and EC will improve our ability to develop effective treatment modalities for EC and will be a vital step toward reducing EC in women globally. DERR1-10.2196/48127.

Heritability of Body Weight, Growth Rate Indices and Their Association with Insulin-Like Growth Factor-2 Gene Polymorphism in Purebred Nigerian Indigenous Chickens and their Crosses with Marshal Chickens

This research was conducted to estimate heritability of body weight and growth rate indices and also to determine their association with insulin-like growth factor-2 genes (IGF-2) polymorphism in purebred Nigerian indigenous chickens and their crosses with Marshal chickens. A total of 300 progenies were generated from the mating of three genotypes of F5 generation of Nigerian indigenous chickens (Normal feather (N), Frizzle feather (F), Naked neck (NN)) and Marshal (M) chickens. Blood samples were collected at twenty weeks for DNA extraction and IGF-2 gene amplification. IGF-2 gene was digested by restriction enzyme (Pst1) and genotyped using restriction fragment length polymorphism method. Growth rate indices (absolute (AGR) and relative growth rates (RGR)) were estimated and associated with IGF-2 gene polymorphism. The results show that IGF-2 allelic variants had no effect on the body weight (BW), AGR and RGR while chicken’s genotype effect was significant (p&lt;0.01) on BW and AGR. Two crossbred genotypes (NxM,1524.23±137.20 and 1894.92±122.81), (FxM, 1445.19±177.24 and 1840.88±177.86) had better performance for AGR and BW, respectively, compared to the purebred of Nigerian indigenous chickens. The heritability values ranged from low (.06±.021) to high (.77±.24).

Effect of dietary intervention with Capsicum annuum extract on growth performance, physiological status, innate immune response, and related gene expression in Nile tilapia

The red pepper (Capsicum annuum) has gained great attention recently because of its biological and pharmacological characteristics. The present approach aimed to evaluate the effects of C. annuum alcoholic extract (CAE) supplementation on Nile tilapia (Oreochromis niloticus) growth performance, physiological status, some metabolic, immune, and regulatory genes expression, and resistance against Streptococcus agalactiae infection. Fish (22.26 ± 0.19 g) were assigned to four treatments (five replicates, each with 10 fish replicate−1) and fed tested diets for 60 days. The experimental diets were supplemented with CAE at 0, 0.4, 0.8, and 1.6 g kg−1, expressed as CAE0, CAE0.4, CAE0.8, and CAE1.6, respectively. The findings exhibited that CAE dietary supplementation improved growth performance, feed utilization, elevated growth hormone level, and digestive enzyme activities (amylase and protease), and lowered leptin hormone in a level-dependent manner. Boosting the mRNA expression of the transporter proteins (solute carrier family 15 member 2 and solute carrier family 26 member 6) and insulin-like growth factor-1 genes with a decrease in the myostatin gene expression was noticed in the CAE-fed groups. The innate immune (serum bactericidal activity %, complement 3, and phagocytic activity %) and antioxidant (glutathione peroxidase and total antioxidant capacity) parameters were significantly (p < 0.05) improved, and the serum malondialdehyde level was significantly decreased by CAE dietary inclusion. A marked upregulation in the mRNA expression of interleukins (il-1β, il-6, il-8, and il-10), transforming growth factor-β, glutathione peroxidase, and glutathione synthetase genes were observed in CAE-fed groups. Dietary CAE decreased the cumulative mortalities after the challenge with S. agalactiae by 20, 13.33, and 10% in CAE0.4, CAE0.8, and CAE1.6, respectively, compared to the control (40%). Overall, dietary supplementation with CAE could improve growth performance and physiological status, and modulate the expression of several regulatory genes in Nile tilapia. The recommended level of CAE is 1.6 g kg−1 to augment growth and health status.

Growth Axis Somatostatin, Growth Hormone Receptor, and Insulin-like Growth Factor-1 Genes Express and Are Affected by the Injection of Exogenous Growth Hormone in Chinemys reevesii.

In this study, to explore the effect of growth hormone changes on the related genes and regulatory roles of the turtle, PCR amplification, real-time fluorescence quantitative analysis, and enzyme cutting technology were used to clone and sequence the somatostatin (SS) gene, growth hormone receptor (GHR), and insulin-like growth factor-1 (IGF-I) sequence of Chinemys reevesii. The effects of human growth hormone on the mRNA expression of growth-axis-related genes SS, GHR, and IGF-1 in different sexes were observed. The study of the SS gene in turtles using real-time fluorescence quantitative PCR showed that the SS gene was mainly expressed in the nervous system and the digestive system, with the highest expression found in the brain, while the GHR gene and the IGF-I gene were expressed in all tissues of Chinemys reevesii. The SS gene was expressed in the brain, pituitary, liver, stomach, and intestine, with the highest expression in the brain and the lowest expression in the liver. Within 4 weeks of the injection of exogenous growth hormone, the expression level of the SS gene in the brain of both sexes first increased and then decreased, showing a parabolic trend, and the expression level of the experimental group was lower than that of the control group. After the injection of growth hormone (GH), the expression of the GHR gene in the liver of both sexes showed a significant increase in the first week, decreasing to the control group level in the second week, and then gradually increasing. Finally, a significant level of difference in the expression of the GHR gene was reached at 3 and 4 weeks. In terms of the IGF-I gene, the changing trend of the expression level in the liver was the same as that of the GHR gene. After the injection of exogenous growth hormone, although the expression of the SS gene increased the inhibition of the secretion of the GHR gene by the Reeves' turtle, exogenous growth hormone could replace the synthesis of GH and GHR, accelerating the growth of the turtle. The experiments showed that the injection of recombinant human growth hormone affects the expression of SS, GHR, and IGF-1 genes, and promotes the growth of the Reeves' turtle.