The aim of the study was to evaluate the influence of the milking phase on somatic cell count (SCC) in milk obtained from the cisternal and alveolar parts of udders of selected Polish Holstein-Friesian cows. The study also assessed the impact of other genetic and environmental factors on SCC variability in cisternal and alveolar milk, including: the individual cow, lactation stage, age of cow, production level, milking speed, fat-to-protein ratio, and milking type. The research included 15 cows of Polish Holstein-Friesian breed at different ages, lactation stages, and with varying daily milk yield. A total of 210 milk observations were conducted, including 105 for 1 min milking and 105 for 8 min milking. The results obtained in the study indicated that milk obtained during two different milking phases exhibited similar SCC levels (F for LOGSCC = 0.79). The average actual SCC in milk produced by 15 cows in 105 observations for 1 min milking was 219,000 cells/mL, while for 8 min milking it was 229,000 cells/mL. The results were inconclusive, suggesting that SCC in cisternal and alveolar milk must be influenced by factors other than the milking phase. The analysis of variance conducted for this purpose provided the basis for stating a highly statistically significant effect of the individual cow (F for LOGSCC = 147.9), lactation stage (F for LOGSCC = 54.64), age of cow (F for LOGSCC = 12.39), daily production level (F for LOGSCC = 34.49), milking speed (F for LOGSCC = 17.56), and fat-to-protein ratio (F for LOGSCC = 22.99) on the variability of characteristics defining SCC in milk. In summary, SCC is characterized by high variability, influenced by a range of environmental and genetic factors such as the individual cow, lactation stage, age of cow, milking speed, and dietary fat-to-protein ratio. The influence of milking phase (1 min or 8 min) and milking type (morning or evening) should be considered inconclusive based on the entire population studied. For half of the cows, SCC in cisternal milk was higher than in alveolar milk, while for the other half, the situation was reversed. Further observations are required to confirm the hypothesis regarding the extent to which cows\' immunological response to bacterial infections is concentrated in the cisternal or alveolar part of the udder under national environmental conditions.

In vitro maturation (IVM) is a promising fertility restoration strategy for patients with nonobstructive azoospermia or for prepubertal boys to obtain fertilizing-competent spermatozoa. However, in vitro spermatogenesis is still not achieved with human immature testicular tissue. Knowledge of various human testicular transcriptional profiles from different developmental periods helps us to better understand the testis development. This scoping review aims to describe the testis development and maturation from the fetal period towards adulthood and to find information to optimize IVM. Research papers related to native and in vitro cultured human testicular cells and single-cell RNA-sequencing (scRNA-seq) were identified and critically reviewed. Special focus was given to gene ontology terms to facilitate the interpretation of the biological function of related genes. The different consecutive maturation states of both the germ and somatic cell lineages were described. ScRNA-seq regularly showed major modifications around 11 years of age to eventually reach the adult state. Different spermatogonial stem cell (SSC) substates were described and scRNA-seq analyses are in favor of a paradigm shift, as the Adark and Apale spermatogonia populations could not distinctly be identified among the different SSC states. Data on the somatic cell lineage are limited, especially for Sertoli cells due technical issues related to cell size. During cell culture, scRNA-seq data showed that undifferentiated SSCs were favored in the presence of an AKT-signaling pathway inhibitor. The involvement of the oxidative phosphorylation pathway depended on the maturational state of the cells. Commonly identified cell signaling pathways during the testis development and maturation highlight factors that can be essential during specific maturation stages in IVM.

Halari donkey breed is one of the indigenous breeds of India and its population is rapidly decreasing. The Jenny milk is more similar to human milk, exhibits a wide range of probiotic diversity and hypo-allergenicity, especially among infants suffering from cow and buffalo milk protein allergy. Some studies indicated low levels of κ-casein fraction of casein protein in donkey milk. The k-casein gene was not amplified from the DNA derived from the milk somatic cells of Halari donkeys. The Halari donkey milk has low somatic cells count. We report the first isolation of DNA from milk somatic cells of Halari donkeys with subsequent characterization of k-casein gene. Through our work, we showed that the milk somatic cells can be used as a non-invasive source for DNA isolation towards molecular studies. It also proved the presence of k-casein gene in Halari donkey milk by its amplification from isolated DNA.

The testis is a complex organ that undergoes extensive developmental changes from the embryonic stage to adulthood. The development of germ cells, which give rise to spermatozoa, is tightly regulated by the surrounding somatic cells.
To better understand the dynamics of these changes, we constructed a transcriptional cell atlas of the testis, integrating single-cell RNA sequencing data from over 26,000 cells across five developmental stages: fetal germ cells, infants, childhood, peri-puberty, and adults. We employed various analytical techniques, including clustering, cell type assignments, identification of differentially expressed genes, pseudotime analysis, weighted gene co-expression network analysis, and evaluation of paracrine cell-cell communication, to comprehensively analyze this transcriptional cell atlas of the testis.
Our analysis revealed remarkable heterogeneity in both somatic and germ cell populations, with the highest diversity observed in Sertoli and Myoid somatic cells, as well as in spermatogonia, spermatocyte, and spermatid germ cells. We also identified key somatic cell genes, including RPL39, RPL10, RPL13A, FTH1, RPS2, and RPL18A, which were highly influential in the weighted gene co-expression network of the testis transcriptional cell atlas and have been previously implicated in male infertility. Additionally, our analysis of paracrine cell-cell communication supported specific ligand-receptor interactions involved in neuroactive, cAMP, and estrogen signaling pathways, which support the crucial role of somatic cells in regulating germ cell development.
Overall, our transcriptional atlas provides a comprehensive view of the cell-to-cell heterogeneity in the testis and identifies key somatic cell genes and pathways that play a central role in male fertility across developmental stages.

Adult stem cells (ASCs) can be cultured with difficulty from most tissues, often requiring chemical or transgenic modification to achieve adequate quantities. We show here that mouse primary fibroblasts, grown in suspension, change from the elongated and flattened morphology observed under standard adherent culture conditions of generating rounded cells with large nuclei and scant cytoplasm and expressing the mesenchymal stem cell (MSC) marker (Sca1; Ly6A) within 24 h. Based on this initial observation, we describe here a suspension culture method that, irrespective of the lineage used, mouse fibroblast or primary human somatic cells (fibroblasts, hepatocytes and keratinocytes), is capable of generating a high yield of cells in spheroid form which display the expression of ASC surface markers, circumventing the anoikis which often occurs at this stage. Moreover, mouse fibroblast-derived spheroids can be differentiated into adipogenic and osteogenic lineages. An analysis of single-cell RNA sequence data in mouse fibroblasts identified eight distinct cell clusters with one in particular comprising approximately 10% of the cells showing high levels of proliferative capacity expressing high levels of genes related to MSCs and self-renewal as well as the extracellular matrix (ECM). We believe the rapid, high-yield generation of proliferative, multi-potent ASC-like cells via the process we term suspension-induced stem cell transition (SIST) could have significant implications for regenerative medicine.

Mastitis is a disease of productive cows, widespread throughout the world, and characterized by significant economic damage to the dairy industry. The subclinical form of this disease is aggravated by additional difficulties with its diagnosis and the lack of clear treatment protocols.
Therefore, the study of the effectiveness of diagnostic studies and the search for new methods of treatment of latent forms of mastitis is an important direction in scientific research in countries with developed dairy cattle breeding.
Studies conducted on the number of dairy cows of the production cooperative \"Izhevsky\" of the Akmola region of the Republic of Kazakhstan showed that when using rapid tests Kenotest, Somatest, Mastidine test, and Wideside test, the same results were obtained when the disease was detected in cows. The effectiveness of the tests was at the level of 60%-62% when using the settling sample as a control. Medical procedures were carried out using the Aquaton-2 microwave radiation apparatus and a homeopathic preparation. When using physiotherapy with microwave radiation, a decrease in the level of microbial contamination of milk from the treated part of the udder by 1.5-5 times was observed.
Biologically active substances of plant origin in the homeopathic preparation, due to the immunostimulating effect, made it possible to increase the level of γ-globulins in the blood serum of sick animals during the application.
The complex use of both methods in the treatment of animals with a subclinical form of mastitis made it possible to reduce the level of somatic cells in the milk of the affected udder lobe to a level that cannot be determined using Kenotest in 4-6 days, which is 2-4 days faster than using these methods separately.

The multicellular green alga Volvox carteri has emerged as a valuable model organism for investigating various aspects of multicellularity and cellular differentiation, photoreception and phototaxis, cell division, biogenesis of the extracellular matrix and morphogenetic movements. While a range of molecular tools and bioinformatics resources have been made available for exploring these topics, the establishment of cell type-specific promoters in V. carteri has not been achieved so far. Therefore, here, we conducted a thorough screening of transcriptome data from RNA sequencing analyses of V. carteri in order to identify potential cell type-specific promoters. Eventually, we chose two putative strong and cell type-specific promoters, with one exhibiting specific expression in reproductive cells (gonidia), the PCY1 promoter, and the other in somatic cells, the PFP promoter. After cloning both promoter regions, they were introduced upstream of a luciferase reporter gene. By using particle bombardment, the DNA constructs were stably integrated into the genome of V. carteri. The results of the expression analyses, which were conducted at both the transcript and protein levels, demonstrated that the two promoters drive cell type-specific expression in their respective target cell types. Transformants with considerably diverse expression levels of the chimeric genes were identifiable. In conclusion, the screening and analysis of transcriptome data from RNA sequencing allowed for the identification of potential cell type-specific promoters in V. carteri. Reporter gene constructs demonstrated the actual usability of two promoters. The investigated PCY1 and PFP promoters were proven to be potent molecular tools for genetic engineering in V. carteri.

BACKGROUND: Most of the resources that support the early development of the embryo are stored in the oocyte. Clearing of maternal resources and activation of the embryonic genome to produce its own mRNA transcripts marks the maternal-to-embryo transition. Dependence on stored mRNA can last from a few hours to several days, depending on animal species. The mechanisms regulating stabilization and recruitment of stored maternal transcripts have not yet been described in full detail but are known to involve reversible polyadenylation and modulation of 3\'UTR-mediated elements. RNA epigenetic modifications, new players in this field, have an important role in RNA regulation and stabilization.
RESULTS: The objectives of this study were first to determine if some of post-transcriptional methylation of stored mRNA is greater in oocytes than in somatic cells. We found that m6A, known to be the most prevalent and involved in various aspects of RNA metabolism and physiological functions, is particularly abundant in porcine oocyte mRNA compared to liver used as a somatic tissue reference. The second objective was to compare the epitranscriptome machinery, such as methyltransferases (\"writers\"), binding proteins (\"readers\") and demethylases (\"erasers\") catalyzing the different process, in follicles and oocytes of different mammalian species by immunofluorescence and confocal microscopy. The expression and localization patterns of these proteins differ between mice, pigs and cows ovaries and oocytes. m5C-associated proteins were generally less abundant. In contrast, m6A-associated proteins were expressed strongly during the early and late stages of folliculogenesis. Transzonal projections were found to contain more granules bearing the m5C mark in mice but both m5C and m6A methylation marks in association with mature oocytes of pigs and cows. Eraser proteins showed the greatest interspecies diversity in terms of distribution in the germinal tissues.
CONCLUSIONS: So far, few studies have looked at the oocyte and ovarian epitranscriptomic profile. Our findings indicate that a hitherto unrecognized species-specific layer of transcript regulation occurs at the RNA level and might be consequential during the oocyte transcriptional silencing period.

Spermatogenesis and testis development are highly structured physiological processes responsible for post-pubertal fertility in stallions. Spermatogenesis comprises spermatocytogenesis, meiosis, and spermiogenesis. Although germ cell degeneration is a continuous process, its effects are more pronounced during spermatocytogenesis and meiosis. The productivity and efficiency of spermatogenesis are directly linked to pubertal development, degenerated germ cell populations, aging, nutrition, and season of the year in stallions. The multiplex interplay of germ cells with somatic cells, endocrine and paracrine factors, growth factors, and signaling molecules contributes to the regulation of spermatogenesis. A cell-to-cell communication within the testes of these factors is a fundamental requirement of normal spermatogenesis. A noteworthy development has been made recently on discovering the effects of different somatic cells including Leydig, Sertoli, and peritubular myoid cells on manipulation the fate of spermatogonial stem cells. In this review, we discuss the self-renewal, differentiation, and apoptotic roles of somatic cells and the relationship between somatic and germ cells during normal spermatogenesis. We also summarize the roles of different growth factors, their paracrine/endocrine/autocrine pathways, and the different cytokines associated with spermatogenesis. Furthermore, we highlight important matters for further studies on the regulation of spermatogenesis. This review presents an insight into the mechanism of spermatogenesis, and helpful in developing better understanding of the functions of somatic cells, particularly in stallions and would offer new research goals for developing curative techniques to address infertility/subfertility in stallions.

Understanding how cooperation evolved and is maintained remains an important and often controversial topic because cheaters that reap the benefits of cooperation without paying the costs can threaten the evolutionary stability of cooperative traits. Cooperation-and especially reproductive altruism-is particularly relevant to the evolution of multicellularity, as somatic cells give up their reproductive potential in order to contribute to the fitness of the newly emerged multicellular individual. Here, we investigated cheating in a simple multicellular species-the green alga Volvox carteri, in the context of the mechanisms that can stabilize reproductive altruism during the early evolution of clonal multicellularity. We found that the benefits cheater mutants can gain in terms of their own reproduction are pre-empted by a cost in survival due to increased sensitivity to stress. This personal cost of cheating reflects the antagonistic pleiotropic effects that the gene coding for reproductive altruism-regA-has at the cell level. Specifically, the expression of regA in somatic cells results in the suppression of their reproduction potential but also confers them with increased resistance to stress. Since regA evolved from a life-history trade-off gene, we suggest that co-opting trade-off genes into cooperative traits can provide a built-in safety system against cheaters in other clonal multicellular lineages.