The sperm protein Spaca6 is essential for fertilization in zebrafish

Fertilization is a key process in all sexually reproducing species, yet the molecular mechanisms that underlie this event remain unclear. To date, only a few proteins have been shown to be essential for sperm-egg binding and fusion in mice, and only some are conserved across vertebrates. One of these conserved, testis-expressed factors is SPACA6, yet its function has not been investigated outside of mammals. Here we show that zebrafish spaca6 encodes for a sperm membrane protein which is essential for fertilization. Zebrafish spaca6 knockout males are sterile. Furthermore, Spaca6-deficient sperm have normal morphology, are motile, and can approach the egg, but fail to bind to the egg and therefore cannot complete fertilization. Interestingly, sperm lacking Spaca6 have decreased levels of another essential and conserved sperm fertility factor, Dcst2, revealing a previously unknown dependence of Dcst2 expression on Spaca6. Together, our results show that zebrafish Spaca6 regulates Dcst2 levels and is required for binding between the sperm membrane and the oolemma. This is in contrast to murine SPACA6, which was reported not to be required for sperm-egg membrane binding but necessary for fusion. These findings demonstrate that Spaca6 is essential for zebrafish fertilization and is a conserved sperm factor in vertebrate reproduction.

Preventive Effect of Egg Membrane Protein Supplementation on Dragon Boat Sports Injury under the Monitoring of Artificial Intelligence and Big Data

Dragon boat is an excellent traditional national sport in China, with excellent fitness results. Based on the country’s strong support for traditional national sports, the dragon boat has developed into a traditional national sport with specific influence and popularity among the masses. Eggshell membrane (ESM) has the effect of relieving arthritis, joint pain, and joint stiffness. The supplementary ESM’s preventive effect on sports injuries has attracted people’s attention. Based on this, this article builds a sports injury prediction model under artificial intelligence and big data monitoring to accurately predict the injuries suffered by athletes, provide help to reduce athletes’ injuries during training, and promote the development of dragon boat sports. This article selects 20 members of a dragon boat team from a university and randomly divides them into a control group (supplement of equal amounts of whey protein), a normal group, an ESM low-dose group (26 mg/kg bw/d), and an ESM high-dose group (52 mg/kg bw/d), with 5 people in each group. The subjects were subjected to downhill exercise and immune stimulation experiments. Among them, the subjects in the control group performed downhill exercise for 12 hours, and they were tested for lactic acid (LAC), lactate dehydrogenase (LDH), urea, malondialdehyde (MDA), hemoglobin (Hb), blood creatinine (CRE), and other related physical and chemical indicators. The other three groups of subjects did not take low-dose and high-dose ESM for 7 consecutive days and were injected with lipopolysaccharide on the 8th day for immune stimulation and tested for interferon (IFN-γ), tumor necrosis factor (TNF-α), interleukins (IL-1β, IL-6, and IL-10), and other related cytokines. The results show that oral egg membrane protein (ESM) can reduce subjects’ serum LAC, LDH, urea, MDA, Hb, and CRE concentration. In addition, the ESM can reduce the expression of proinflammatory factors IL-1β and TNF-α, while increasing the expression of anti-inflammatory factor IL-10, which provides clinical data for the clinical use of ESM as a supplement for sports injury or for relieving arthritis symptoms. The study shows that egg membrane protein has a protective effect against sports injury and may be regulated by inflammatory factors.

Synchrotron imagery of phosphatized eggs in Waptia cf. W. fieldensis from the middle Cambrian (Miaolingian, Wuliuan) Spence Shale of Utah

Exceptionally preserved fossil eggs and embryos provide critical information regarding paleoembryogenesis, reproductive strategies, and the early ontogeny of early arthropods, but the rarity of preservation of both eggs and egg-bearing organisms in situ limits their use in detailed evolutionary developmental (evo-devo) studies. Burgess Shale-type deposits preserve rare instances of egg-bearing arthropods as carbonaceous compressions; however, the eggs are usually poorly preserved with no compelling evidence of embryos. We describe the first record of a brooding specimen of Waptia cf. W. fieldensis from the Spence Shale, a Cambrian (Wuliuan Stage) Burgess Shale-type deposit in northeastern Utah and southeastern Idaho. This is the first record of an egg-bearing arthropod from the Spence Shale and it exhibits two distinct modes of preservation among eggs within the single clutch: carbonization and phosphatization. Unlike the egg-bearing Burgess Shale specimens, many eggs of this Utah specimen are also preserved three-dimensionally. In addition, synchrotron radiation X-ray tomographic microscopy reveals internal distributions of mineral phases, along with potential remnants of the egg membrane and attachment structures, but, as in the Burgess Shale, no explicit traces of developing embryos. The distinct modes of preservation highlight the existence of diagenetic microenvironments within some eggs, but not in others during fossilization.

Formulation and Evaluation of Ethosomal Nanoparticles of Avobenzone for Improved Sunscreen Efficacy

Ethosomes entrapping avobenzone were prepared using cold method and the effect of varying concentration of ethanol was considered for obtaining an optimized formulation. Lecithin (2%w/w) was used as the phospholipid to provide the structure to the vesicles and propylene glycol (10%) was used as the permeating agent. The vesicles were found to be of spherical to irregular shape ranged from 1.11 µm to 1.6 µm in size. The drug entrapment in the ethosomes was studied by analyzing the unentrapped drug spectrophotometrically. The in vitro permeation study suggested that the maximum permeation in the egg membrane occurred in AET3 (0.40 mg/cm2) with 30% ethanol concentration. It was observed that only about 2% degradation occurred at room temperature and all formulations were almost stable at 8° and 4° with only 1.3% degradation of avobenzone thereby proving the stability of the developed system. The best ethosomal formulation (AET3) was incorporated into gel base to obtain sunscreen gels and the results revealed a good protection of the ethosomal gel when 2% carbopol was used as the gelling base. It could be concluded that incorporation of avobenzone in the ethosomal carrier and formulating the same as gel formulation might help in reducing the dose of avobenzone as well as improving the sunscreen efficacy (sun protection over enhanced duration).

RanGTP and the actin cytoskeleton keep paternal and maternal chromosomes apart during fertilization

Zygotes require two accurate sets of parental chromosomes, one each from the mother and the father, to undergo normal embryogenesis. However, upon egg–sperm fusion in vertebrates, the zygote has three sets of chromosomes, one from the sperm and two from the egg. The zygote therefore eliminates one set of maternal chromosomes (but not the paternal chromosomes) into the polar body through meiosis, but how the paternal chromosomes are protected from maternal meiosis has been unclear. Here we report that RanGTP and F-actin dynamics prevent egg–sperm fusion in proximity to maternal chromosomes. RanGTP prevents the localization of Juno and CD9, egg membrane proteins that mediate sperm fusion, at the cell surface in proximity to maternal chromosomes. Following egg–sperm fusion, F-actin keeps paternal chromosomes away from maternal chromosomes. Disruption of these mechanisms causes the elimination of paternal chromosomes during maternal meiosis. This study reveals a novel critical mechanism that prevents aneuploidy in zygotes.

Experimental Urolithiasis Model to assess Phyto-fractions as Anti-lithiatic Contributors: A Herbaceutical Approach

Life-style disorders have bought a serious burden on the maintenance of health in animals and humans. Lithiasis specifically nephro- and urolithiasis is no exception and needs urgent attention. Currently, only semi-invasive and surgical methods are widely employed which leads to trauma and reoccurrence of kidney stones. Hence complementary and alternative herbal medicine could pave newer ways in exploring anti-lithiatic contributors. The current study attempts to screen twenty herbal hot aqueous leaf extracts for assessing their antioxidant potency (anti-stress) and efficiency against urolithiasis in an experimental calcium oxalate-induced in vitro (chicken egg membrane) model. The study was further validated by In silico molecular docking studies using the Molegro software package on enzymatic biomarkers involved in scavenging oxidants in the host and regulating oxalate metabolism at a cellular level. Among the screened botanicals Kalanchoe pinnata exhibited promising results compared to the standard chemical (potassium-magnesium citrate) and phyto-formulation drug (cystone) currently used by clinicians for treating urolithiasis. The phytochemical profiling (qualitative and quantitative) and virtual studies indicated rutin from Kalanchoe pinnata as a potential candidate for preventing kidney stones. The results of the current study provide better insights into the design and development of newer, smart, and cost-effective herbal therapeutics making food as medicine.

Spinochrome Identification and Quantification in Pacific Sea Urchin Shells, Coelomic Fluid and Eggs Using HPLC-DAD-MS

The high-performance liquid chromatography method coupled with diode array and mass spectrometric detector (HPLC-DAD-MS) method for quinonoid pigment identification and quantification in sea urchin samples was developed and validated. The composition and quantitative ratio of the quinonoid pigments of the shells of 16 species of sea urchins, collected in the temperate (Sea of Japan) and tropical (South-China Sea) climatic zones of the Pacific Ocean over several years, were studied. The compositions of the quinonoid pigments of sea urchins Maretia planulata, Scaphechinus griseus, Laganum decagonale and Phyllacanthus imperialis were studied for the first time. A study of the composition of the quinonoid pigments of the coelomic fluid of ten species of sea urchins was conducted. The composition of quinonoid pigments of Echinarachnius parma jelly-like egg membrane, of Scaphechinus mirabilis developing embryos and pluteus, was reported for the first time. In the case of Scaphechinus mirabilis, we have shown that the compositions of pigment granules of the shell epidermis, coelomic fluid, egg membrane, developing embryos and pluteus are different, which should enable a fuller understanding of the functions of pigments at different stages of life.

Zebrafish stm is involved in the development of otoliths and of the fertilization envelope

Using an in vivo assay, we selected 11 genes that were highly upregulated during the induction of ovulation in zebrafish using microarray analysis and RNA sequencing. The starmaker gene (stm) was one of these genes. Although stm has been previously reported to be involved in otolith formation during the early development of zebrafish, we detected its expression in eggs and showed that stm was related to fertilization by establishing an stm gene knockout strain using the CRISPR/Cas9 system. Further phenotypic analysis of stm knockout fish was conducted in this study. With a higher nonfertilization rate, the stm mutant strain showed an extremely low survival rate. Otoliths of stm homozygous mutant zebrafish showed abnormal morphology in embryos and adult fish. However, fish did not show any abnormalities in swimming behaviour in either embryos or adults. Stm proteins were detected on the chorion of ovulated eggs before spawning. Fibre-supported knob-like structures on the fertilization envelope (FE) also showed abnormal structures in stm mutants. The Stm protein is necessary for otolith formation, and a lack of Stm causes abnormal otolith formation. The partial defect of otolith formation does not cause defects in swimming behaviour. The Stm protein is expressed in the chorion and is responsible for the formation of fibre-supported knob-like structures on the FE. It was suggested that a lack of Stm caused a lower fertilization rate due to inadequate formation of the FE. Lay summary In zebrafish, the protein Starmaker (Stm) was identified as having a role in ovulation. Stm is also known to be required for the formation of ear stones (otoliths) which are needed to keep the body in balance. Zebrafish lacking Stm were produced by genome editing. As expected, Stm-deficient fish formed abnormal otoliths. To investigate the role of Stm in ovulation, fertilization and early development, we tried mating of Stm mutants and observed their juveniles. Although no problem found in ovulation, we found low fertilization rate and abnormal structure of knob-like structure (small pit) on the egg membrane. Survival rate of embryos with abnormal egg membrane was extremely low. It was demonstrated that Stm protein is necessary to form the functional egg membrane to protect embryos from the outside environment.