Ultrastructural Morphology Research Articles

Development of Transethosomes Delivery System Loaded with Bouea macrophylla Griffith Seed Kernel Extract for Cosmeceutical Application

This research aimed to develop and optimize transethosomes loaded with Bouea macrophylla Griffith seed kernel extract (B-SE). Firstly, extraction procedures using solvent fractionation with various polarity: hexane, ethyl acetate and 95% (v/v) ethanol. The physicochemical properties of B-SE were investigated in terms of partition coefficient (LogP), solubility and pH-stability. To optimize the transethosomes formulation, the compositions were varied in sorts of percent cosurfactant, types of cosurfactant, extract concentration, percent total surfactant and types of particle ingredients (phospholipid or cholesterol). The significant factors influencing physical characteristics of the transethosomes were statistically identified using the 25-1 fractional factorial design. Particle size, polydispersity index (PDI), percent entrapment efficiency (%EE) and ultrastructural morphology of the optimized formulation was then characterized. As a marker of B-SE, ellagic acid (EA) was identified by HPLC at a retention time of 32.506 ± 0.16 min. The LogP of B-SE was 0.17. B-SE was practically insoluble in water but freely soluble in propylene glycol and ethanol. The pH condition that stabilized the extract was pH-5. The optimized formulation, consisted of phospholipid, propylene glycol, TWEEN 20, SPAN 80 and 2.5 mg/ml of B-SE presenting particle sizes of 79.72 ± 2.42 nm, PDI of 0.26±0.02 and %EE of 83.10±0.03% along with a spherical particle shape. To conclude, the optimized transethosomes loaded with B-SE fabricated by phospholipid and propylene glycol has desirable attributes for further developing into an anti-acne cosmeceutical. Nonetheless, the bioactivities and safety profile of the transethosomes were necessary for further investigation. Keywords: Bouea macrophylla Griffith, Transethosomes, Ellagic acid, Fractional factorial design, Nanocarrier

Antimicrobial mechanisms of g-C3 N4 @ZnO against oomycetes Phytophthora capsici: from its metabolism, membrane structures and growth.

Phytophthora capsici, a refractory and model oomycete plant pathogen, especially threatens multiple vegetable crops. A limited number of chemical pesticides play a vital role in controlling oomycete plant diseases. However, this approach often leads to excessive use of chemical agent, exacerbates environmental issues and more and more drug-resistant strains of oomycete. Therefore, it is imperative to devise innovative solutions that can effectively address the infection of oomycete while maintaining high levels of environmental sustainability and low toxicity. In this study, g-C3 N4 @ZnO heterostructure was synthesized and characterized. The g-C3 N4 @ZnO showed higher toxicity on Phytophthora capsici than graphitic carbon nitride (g-C3 N4 ) nanosheets and zinc oxide (ZnO) nanoparticles in vitro and in vivo. Except the hyphal growth of Phytophthora capsici, their germination rate of spores, sporangium formation and number of spores were all suppressed by g-C3 N4 @ZnO heterostructure. Furthermore, we found that this g-C3 N4 @ZnO heterostructure has higher photocatalytic activity under visible light, which potentially enhanced the reactive oxygen species (ROS) mediated stress on Phytophthora capsici. Ultrastructural morphology, global changes of gene expression and weighted gene co-expression network analysis all supported that the anti-oomycete activity of g-C3 N4 @ZnO was manifested in the destruction of membrane system and inhibition of multiple metabolisms of Phytophthora capsici under visible irradiation, which also could be attributed to the ROS and zinc ion (Zn2+ ) mediated stress. This works offers a novel oomycete disease management strategy by using g-C3 N4 @ZnO, which were attributed to the ROS stress, destruction of membrane system and inhibition of multiple metabolisms. © 2023 Society of Chemical Industry.

The ultrastructure of nymphal dermal pores and the genitalia of adult male of Phenacoccus solenopsis (Hemiptera: Pseudococcidae)

The cotton mealybug, Phenacoccus solenopsis, has established itself as an invasive insect pest worldwide. It causes structural and physiological damage to various crops and can cause substantial financial losses in their production. The successful reproduction of this pest under a wide range of conditions is a key to its success. Despite this, the morphology of its genitalia, genital sensilla, and wax-producing dermal pores has received little attention, with little descriptions of their ultrastructure. By investigating those features with SEM, the present study revealed considerable new insights into the identification of the nymphal and adult stages of P. solenopsis. In addition, the description of the ultrastructural genital morphology of the immature stages of P. solenopsis has revealed characteristics that facilitate their discrimination. Trilocular pores were observed on both sides of the body, while the quinquelocular pores were distributed only on the ventral surface in both the first and second nymphal instars. The adult male is characterized by two pairs of waxy caudal filaments surrounded by clusters of 55 to 60 stellate pores, and each pregenital segment bears a pair of stellate pores composed of 4 or 5 peripheral loculi. Sensilla trichodea and numerous microtrichia are present on the pregenital segments. The penile sheath bears three subtypes of sensilla basiconica and also campaniformia, whereas the style bears three subtypes of sensilla campaniformia. The findings of this study could assist in the identification of the adult and nymphal stages of P. solenopsis, and also provide insights into the structures found on the genitalia of the adult male that possibly have an important role in mating events and copulatory behavior. Furthermore, these findings were able to contribute to better understanding the functional morphology of P. solenopsis.

Infant glycogen storage disease type Ⅳ: a clinicopathological and genetic characteristics analysis of five cases

Objective: To investigate the clinical pathology and gene mutation characteristics of patients with glycogen storage disease type Ⅳ (GSD Ⅳ). Methods: The clinical data, liver histopathology and ultrastructural morphology, and gene sequencing results of 5 GSD Ⅳ cases diagnosed in the Children's Hospital Affiliated to Shanghai Jiaotong University School of Medicine and the Children's Hospital of Fudan University from January 2015 to February 2022 were collected and analyzed retrospectively. Results: Among the 5 cases, 3 were male and 2 were female, ranging in age from 4 months to 1 year and 9 months. The clinical manifestations included fever, hepatosplenomegaly, liver insufficiency, growth retardation and hypotonia. Four cases had liver biopsy showing ground-glass-like changes in hepatocytes with intracytoplasmic inclusion bodies and varying degrees of fibrosis. Liver electron microscopy in 2 cases showed that the level of glycogen increased to varying degrees, and the cytoplasm was filled with low electron density substances. Genetic testing revealed that 3 cases had compound heterozygous variants in GBE1 gene; 1 case had a single pathogenic variant in GBE1 gene; and 1 case was deceased with no genetic testing, but each parent was tested for a heterozygous variant in the GBE1 gene. A total of 9 GBE1 gene mutations were detected, 3 of which were reported mutations and 6 novel mutations. One case died of liver cirrhosis, and 1 case underwent autologous liver transplantation. After transplantation, the liver function basically returned to normal, and the growth and development improved; the other 3 cases were managed through diet control and symptomatic treatment. Conclusions: CSD Ⅳ is an extremely rare inherited metabolic disease caused by GBE1 gene mutation, often presenting with hepatic and neuromuscular disorders, with heterogeneous clinical manifestations. The diagnosis mainly depends on histopathology and a pedigree gene analysis.

α-Mangostin and lawsone methyl ether in tooth gel synergistically increase its antimicrobial and antibiofilm formation effects in vitro

Objectivesα-Mangostin (α-MG) and lawsone methyl ether (LME) show antimicrobial and anti-biofilm activities. The objectives of this study were to develop a herbal tooth gel containing α-MG and LME plus fluoride and determine its antimicrobial, anti-biofilm formation, anti-cancer, anti-inflammatory, wound healing, and enamel microhardness effects.MethodsAntimicrobial assays against Streptococcus mutans, Porphyromonas gingivalis, and Candida albicans were performed. The microbes’ ultrastructural morphology was assessed using Transmission Electron Microscopy. The effect on microbial biofilm formation was tested by a broth microdilution. Cell viability was assessed with MTT assay. The anti-inflammatory effect was investigated by measuring inhibition of nitric oxide production. Enamel microhardness was measured via Vickers microhardness testing. The enamel chemical composition was investigated with Fourier Transform Spectrometer. The enamel surface morphology and fluoride content were examined by Scanning Electron Microscopy and Energy Dispersive X-ray Spectroscopy.ResultsThe results show synergistic effects of α-MG and LME on antimicrobial activity and antibiofilm formation without cytotoxicity at a therapeutic dose. At a higher dose, the tooth gel inhibited proliferation of cancer cell line. Enamel microhardness was increased after brushing with the tooth gel plus fluoride. A large amount of fluoride was detected on the enamel surface.ConclusionThe tooth gel containing α-MG and LME synergized its antimicrobial activity and antibiofilm formation and inhibited oral cancer cell proliferation. Incorporating fluoride into the tooth gel increased enamel microhardness. Thus, the herbal tooth gel containing α-MG and LME plus fluoride may be useful for preventing dental caries and promoting oral health.

Effects of habitats in typical karst areas of Guizhou on ultrastructural morphology of Typhlocybinae.

Biology and the environment are inextricably linked. Insects are intricately linked to their habitats as part of the ecosystem. In this study, scanning electron microscopy (SEM) observations showed two sensilla chaetica, several sensilla trichodea, and at least one sensilla basiconicum on the antennae of typhlocybine insects. There were no sensilla on the labrum; however, the surface morphology was different. There were more sensilla trichodea on the surface, mostly symmetrically distributed along the labial groove, and there was little difference in the structure of the stylet fascicle. The correlation between the sensilla number on the body surface of typhlocybine insects and environmental factors in the 3 study areas was as follows: Huajiang > Bijie > Shibing, which is consistent with the classification of rocky desertification grade; that is, the higher the rocky desertification grade, the greater the sensilla number on leafhoppers affected by the environment. The correlation between the number of leafhoppers and environmental factors in the 3 months was as follows: end of September > end of May > end of July, which was consistent with the changing temperature trends. The results of this study enrich our knowledge of the morphological characteristics of leafhoppers and explore the potential value of insect surface ultra-morphology for use by humans.

Synergistic effect of commercial mangosteen extract (Garcinia mangostana L.) and amoxicillin against methicillin-resistant Staphylococcus aureus (MRSA)

Antibiotic resistance occurs worldwide, and has become a threat to humankind. Previous data have shown that antimicrobial resistance is a global issue demanding immediate resolution because it threatens the environment and society. The present work thus investigated the synergistic effects of commercial Garcinia mangostana L. (GML) extract and amoxicillin on the growth of methicillin-resistant Staphylococcus aureus (MRSA) bacterial cells. A commercial GML extract was screened for phytochemical properties, and the presence of α-mangostin was detected using high-performance liquid chromatography (HPLC). The antibacterial activity of the commercial GML extract with amoxicillin was analysed by minimum inhibitory concentration (MIC) and checkerboard assays. The morphology ultrastructure of bacteria was observed using transmission electron microscopy (TEM), after treatment with commercial GML extract, either single or in combination with amoxicillin. The MICs of amoxicillin and commercial GML extract against MRSA bacteria were 250.00 and 137.50 μg/mL, respectively. The checkerboard assay showed synergistic activity in the combination of commercial GML extract (34.38 µg/mL) and amoxicillin (62.50 µg/mL) at fractional inhibitory concentration (FIC) index of < 0.5. Damage to the structure of bacteria occurred due to the commercial GML extract plus amoxicillin. It was observed that the loss of bacterial cell membranes led to an irregular bacterial structure. These findings provided evidence that the combination of commercial GML extract and amoxicillin could reverse bacterial resistance in order to determine the susceptibility of traditional drugs.

Preclinical assessment of IL-1β primed human umbilical cord mesenchymal stem cells for tendon functional repair through TGF-β/IL-10 signaling

BackgroundInadequate repair capacity and disturbed immune compartments are the main pathological causes of tendinopathy. Transplantation of mesenchymal stem cells (MSCs) become an effective clinic option to alleviate tendinopathy. Interleukin-1β (IL-1β) could confer on MSCs enhanced immunoregulatory capability to remodel the repair microenvironment favoring tissue repair. Therefore, IL-1β activated UC-MSCs (1βUC-MSCs) may exert favorable efficacy in promoting tendon repair in a preclinical tendinopathy rat model. MethodsTendon-derived stem cells (TDSCs) were isolated and characterized. In vitro, the levels of immunoregulatory-related cytokines such as IL-1β, IL-6, IL-10, and TGF-β secreted by 1βUC-MSCs and unprimed UC-MSCs was measured. And tendon-specific markers expressed by TDSCs cultured with primed cultured medium (CM) or unprimed CM were detected. In vivo, Achilles tendinopathy was induced by 30 μL collagenase I injection in Sprague Dawley rats. One week later, the rats were randomly injected with UC-MSCs primed with IL-1β (106 cells per tendon), UC-MSCs, or PBS. After rats were sacrificed, histological evaluation, electron microscopy, biomechanical tests, gait performance were conducted to evaluate the structural and functional recovery of Achilles tendons. The inflammation and metabolic state of the extracellular matrix, and the potential mechanism were assessed by immunohistochemical staining and Western blot. ResultsUC-MSCs were activated by IL-1β to secrete higher levels of IL-10 and TGF-β while the secretion levels of IL-6 and IL-1β were not changed significantly, promoting a higher expression level of COL I and TNMD in TDSCs under proinflammatory environment. In vivo, the transplanted 1βUC-MSCs could survive up to 5 weeks after injection with tenogenic differentiation and improved tendon healing histologically semi-quantified by modified Bonar scores. This structural regeneration was further confirmed by observation of ultrastructural morphology, and led to good functional recovery including improved biomechanical properties and gait performance. During this process, the inflammatory response and metabolism of the extracellular matrix was improved through TGF-β/IL-10 pathway. ConclusionThis study demonstrated that the transplantation of UC-MSCs activated by IL-1β exhibited satisfactory ability for promoting tendon functional repair in a tendinopathy rat model. During this process, the balance of inflammatory response and extracellular matrix metabolism was remodeled, and the TGF-β/Smad2/3 and IL-10 signaling pathways were activated simultaneously. We cautiously conclude that the IL-1β primed UC-MSCs could be a promising strategy for enhancing the ability of MSCs to treat tendinopathy.

Fibrocystin/Polyductin releases a C-terminal fragment that translocates into mitochondria and suppresses cystogenesis

Fibrocystin/Polyductin (FPC), encoded by PKHD1, is associated with autosomal recessive polycystic kidney disease (ARPKD), yet its precise role in cystogenesis remains unclear. Here we show that FPC undergoes complex proteolytic processing in developing kidneys, generating three soluble C-terminal fragments (ICDs). Notably, ICD15, contains a novel mitochondrial targeting sequence at its N-terminus, facilitating its translocation into mitochondria. This enhances mitochondrial respiration in renal epithelial cells, partially restoring impaired mitochondrial function caused by FPC loss. FPC inactivation leads to abnormal ultrastructural morphology of mitochondria in kidney tubules without cyst formation. Moreover, FPC inactivation significantly exacerbates renal cystogenesis and triggers severe pancreatic cystogenesis in a Pkd1 mouse mutant Pkd1V/V in which cleavage of Pkd1-encoded Polycystin-1 at the GPCR Proteolysis Site is blocked. Deleting ICD15 enhances renal cystogenesis without inducing pancreatic cysts in Pkd1V/V mice. These findings reveal a direct link between FPC and a mitochondrial pathway through ICD15 cleavage, crucial for cystogenesis mechanisms.

HivNet: Studying in Depth the Morphology of HIV-1 Virion Using Deep Learning

Purpose: Human Immunodeficiency Virus (HIV) continues to be a disease that kills thousands of individuals each year. The HIV infection is incurable. However, HIV infection has turned into a treatable chronic health condition because of improved access to efficient HIV prevention, diagnosis, treatment, and care. Transmission Electron Microscopy’s (TEM) ability to directly visualize virus particles and distinguish ultrastructure morphology at the nanometer scale, makes it useful in HIV-1 research where it is used for assessing the actions of inhibitors that obstruct the maturation and morphogenesis phases of the virus lifecycle. Hence with its use, the disease's serious stage can be avoided by receiving an early diagnosis.
 Materials and Methods: Through the dedicated use of computer vision frameworks and machine learning techniques, we have developed an optimized low-computational-cost 8-layer Convolutional Neural Network (CNN) backbone capable of classifying HIV-1 virions at various stages of maturity and morphogenesis. The dataset including TEM images of HIV-1 viral life cycle phases is analysed and augmented through various techniques to make the framework robust in real-time. The CNN layers then extract pertinent disease traits from TEM images and utilise them to provide diagnostic predictions.
 Results: It was discovered that the framework performed with an accuracy of 99.76% on the training set, 85.83% on the validation set, and 91.33% on the test set, after being trained on a wide range of micrographs which comprised of different experimental samples and magnifications.
 Conclusion: The suggested network's performance was compared to that of other state-of-the-art networks, and it was discovered that the proposed model was undisputed for classifying TEM images of unseen HIV-1 virion and required less time to train and tweak its weights. The framework can operate more effectively than machine learning algorithms that consume a lot of resources and can be deployed with limited computation and memory resource requirements.

Characterization of the extracellular vesicles, ultrastructural morphology, and intercellular interactions of multiple clinical isolates of the brain-eating amoeba, Naegleria fowleri.

As global temperatures rise to unprecedented historic levels, so too do the latitudes of habitable niches for the pathogenic free-living amoeba, Naegleria fowleri. This opportunistic parasite causes a rare, but >97% fatal, neurological infection called primary amoebic meningoencephalitis. Despite its lethality, this parasite remains one of the most neglected and understudied parasitic protozoans. To better understand amoeboid intercellular communication, we elucidate the structure, proteome, and potential secretion mechanisms of amoeba-derived extracellular vesicles (EVs), which are membrane-bound communication apparatuses that relay messages and can be used as biomarkers for diagnostics in various diseases. Herein we propose that N. fowleri secretes EVs in clusters from the plasma membrane, from multivesicular bodies, and via beading of thin filaments extruding from the membrane. Uptake assays demonstrate that EVs are taken up by other amoebae and mammalian cells, and we observed a real-time increase in metabolic activity for mammalian cells exposed to EVs from amoebae. Proteomic analysis revealed >2,000 proteins within the N. fowleri-secreted EVs, providing targets for the development of diagnostics or therapeutics. Our work expands the knowledge of intercellular interactions among these amoebae and subsequently deepens the understanding of the mechanistic basis of PAM.

Investigation of conditioned medium properties obtained from human umbilical cord mesenchymal stem/stromal cells preconditioned with dimethyloxalylglycine in a correlation with ultrastructural changes.

Mesenchymal stem/stromal cells (MSCs) hold significant therapeutic value due to their regeneration abilities, migration capacity, and immunosuppressive and immunomodulatory properties. These cells secrete soluble and insoluble factors, and this complex secretome contributes to their therapeutic effect. Furthermore, stimulation of cells by various external stimuli lead to secretome modifications that can increase the therapeutic efficacy. So, this study examined the effect of dimethyloxalylglycine (DMOG), a hypoxia-mimetic agent, on secretome profiles and exosome secretions of MSCs by evaluating conditioned medium (CM) and ultrastructural morphologies of the cells in comparison with unpreconditioned MSCs. The appropriate dose and duration of the use of DMOG were determined as 1000 μM and 24 h by evaluating the HIF-1α expression. DMOG-CM and N-CM were collected from MSCs incubated in serum-free medium with/without DMOG for 24 h, respectively. The content analysis of conditioned mediums (CMs) revealed that VEGF, NGF, and IL-4 levels were increased in DMOG-CM. Subsequently, exosomes were isolated from the CMs and were shown by transmission electron microscopy and Western blot analysis in both groups. The effects of CMs on proliferation and migration were determined by in vitro wound healing tests; both CMs increased the fibroblast's migratory and proliferative capacities. According to the ultrastructural evaluation, autophagosome, autolysosome, myelin figure, and microvesicular body structures were abundant in DMOG-preconditioned MSCs. Consistent with the high number of autophagic vacuoles, Beclin-1 expression was increased in those cells. These findings suggested that DMOG could alter MSCs' secretion profile, modify their ultrastructural morphology accordingly, and make the CM a more potent therapeutic tool. RESEARCH HIGHLIGHTS: Preconditioning mesenchymal stem/stromal cells with dimethyloxalylglycine, a hypoxia-mimetic agent, could modify cellular metabolism. Hypoxic mechanisms lead to alterations in the ultrastructural characteristics of mesenchymal stromal/stem cells. Preconditioning with dimethyloxalylglycine leads to ultrastructural and metabolic changes of mesenchymal stromal/stem cells along with modifications in their secretome profiles. Preconditioning of mesenchymal stromal/stem cells could render them a more potent therapeutic tool.

Ultrastructural Changes and Inflammatory Processes of Day-Dependent Cisplatin Administration on Rat Cardiac Tissue

Aim: Cisplatin (CP) is used to treat a variety of cancers as a chemotherapeutic agent. This drug has also severe side effects and its use exhibits serious toxicity in a number of organs, including kidney and heart. The aim of the present study was to evaluate the ultrastructural and inflammatory changes induced by CP treatment in rat cardiac tissue in a time-dependent manner.
 Material and Methods: Rats were randomly divided into three experimental groups; control (only saline), CP D2 (treated with CP 2.5 mg/kg/day for 2 days), and CP D7 (treated with CP 2.5 mg/kg/day for 7 days). Cardiac tissues were examined under an electron microscope. Inflammation markers including tumor necrosis factor-α (TNF-α) and interleukin 1β (IL-1β) were analyzed by immunohistochemistry. In addition, electrocardiography was performed to measure the electrical activity.
 Results: The ultrastructural analysis of the CP D7 group revealed that myofibrils were disrupted and disorganized, mitochondria degenerated, and interstitial edema developed. When compared to the control and CP D2 groups, there was a noticeable increase in the level of TNF-α and IL-1β expression in the CP D7 group according to immunohistochemistry results. Electrocardiography showed that RR interval was longer in CP D7 than CP D2 and control groups.
 Conclusion: CP for 7 days damaged the ultrastructural morphology in cardiac tissue. Therefore, these findings suggest that the potential therapeutic approaches to reduce mitochondrial damage and inflammation against toxicity caused by CP may provide for clinically significant prevention when using the drug for an extended period of time.

The ultrastructural nature of human oocytes’ cytoplasmic abnormalities and the role of cytoskeleton dysfunction

To investigate the structural bases of human oocytes' cytoplasmic abnormalities and the causative mechanism of their emergence. Knowledge of an abnormal oocyte's intracellular organization is vital to establishing reliable criteria for clinical evaluation of oocyte morphology. Laboratory-based study on experimental material provided by a private assisted reproduction clinic. University laboratory and imaging center. A total of 105 women undergoing hormonal stimulation for invitro fertilization (IVF) donated their spare oocytes for this study. Transmission electron microscopy (TEM) was used to analyze the fine morphology of 22 dysmorphic IVF oocytes exhibiting different types of cytoplasmic irregularities, namely, refractile bodies; centrally located cytoplasmic granularity (CLCG); smooth endoplasmic reticulum (SER) disc; and vacuoles. A total of 133 immature oocytes were exposed to cytoskeleton-targeting compounds or matured in control conditions, and their morphology was examined using fluorescent and electron microscopy. The ultrastructural morphology of dysmorphic oocytes was analyzed. Drug-treated oocytes had their maturation efficiency, chromosome-microtubule configurations, and fine intracellular morphology examined. TEM revealed ultrastructural characteristics of common oocyte aberrations and indicated that excessive organelle clustering was the underlying cause of 2 of the studied morphotypes. Inhibition experiments showed that disruption of actin, not microtubules, allows for inordinate aggregation of subcellular structures, resembling the ultrastructural pattern seen in morphologically abnormal oocytes retrieved in IVF cycles. These results imply that actin serves as a regulator of organelle distribution during human oocyte maturation. The ultrastructural analogy between dysmorphic oocytes and oocytes, in which actin network integrity was perturbed, suggests that dysfunction of the actin cytoskeleton might be implicated in generating common cytoplasmic aberrations. Knowledge of human oocytes' inner workings and the origin of morphological abnormalities is a step forward to a more objective oocyte quality assessment in IVF practice.

Contribution of trehalose to ethanol stress tolerance of Wickerhamomyces anomalus

BackgroundThe ascomycetous heterothallic yeast Wickerhamomyces anomalus (WA) has received considerable attention and has been widely reported in the winemaking industry for its distinctive physiological traits and metabolic attributes. An increased concentration of ethanol during ethanol fermentation, however, causes ethanol stress (ES) on the yeast cells. Trehalose has been implicated in improving survival under various stress conditions in microorganisms. Herein, we determined the effects of trehalose supplementation on the survival, differentially expressed genes (DEGs), cellular morphology, and oxidative stress tolerance of WA in response to ES.ResultsThe results indicated that trehalose improved the survival and anomalous surface and ultrastructural morphology of WA. Additionally, trehalose improved redox homeostasis by reducing the levels of reactive oxygen species (ROS) and inducing the activities of antioxidant enzymes. In addition, DEGs affected by the application of trehalose were enriched in these categories including in gene expression, protein synthesis, energy metabolism, and cell cycle pathways. Additionally, trehalose increased the content of intracellular malondialdehyde (MDA) and adenosine triphosphate.ConclusionsThese results reveal the protective role of trehalose in ES mitigation and strengthen the possible uses of WA in the wine fermentation sector.

Pimobendan prevents cardiac dysfunction, mitigates cardiac mitochondrial dysfunction, and preserves myocyte ultrastructure in a rat model of mitral regurgitation

BackgroundPimobendan has been proven to delay the onset of congestive heart failure (CHF) in dogs with mitral regurgitation (MR); however, molecular underlying mechanisms have not been fully elucidated. This study aimed to investigate (1) the effects of pimobendan on cardiac function, cardiac mitochondrial quality and morphology, and cardiac ultrastructure in a rat model of chronic MR and (2) the direct effect of pimobendan on intracellular reactive oxygen species (ROS) production in cardiac cells. MR was surgically induced in 20 Sprague-Dawley rats, and sham procedures were performed on 10 rats. Eight weeks post-surgery, the MR rats were randomly divided into two groups: the MR group and the MR + pimobendan group. Pimobendan (0.15 mg/kg) was administered twice a day via oral gavage for 4 weeks, whereas the sham and MR groups received equivalent volumes of drinking water. Echocardiography was performed at baseline (8 weeks post-surgery) and at the end of the study (4 weeks after treatment). At the end of the study protocol, all rats were euthanized, and their hearts were immediately collected, weighed, and used for transmission electron microscopy and mitochondrial quality assessments. To evaluate the role of pimobendan on intracellular ROS production, preventive or scavenging properties were tested with H2O2-induced ROS generation in rat cardiac myoblasts (H9c2).ResultsPimobendan preserved cardiac functions and structure in MR rats. In addition, pimobendan significantly improved mitochondrial quality by attenuating ROS production and depolarization (P < 0.05). The cardiac ultrastructure and mitochondrial morphology were significantly preserved in the MR + pimobendan group. In addition, pimobendan appeared to play as a ROS scavenger, but not as a ROS preventer, in H2O2-induced ROS production in H9c2 cells.ConclusionsPimobendan demonstrated cardioprotective effects on cardiac function and ultrastructure by preserving mitochondrial quality and acted as an ROS scavenger in a rat model of MR.

SARS-CoV-2–Induced Vasculitic Skin Lesions Are Associated with Massive Spike Protein Depositions in Autophagosomes

In patients infected with severe acute respiratory syndrome coronavirus 2, vasculopathic changes of the skin are associated with a severe prognosis. However, the pathogenesis of this vasculopathy is not conclusively clarified. In this study, 25 prospectively collected skin samples from patients with COVID-19-related skin lesions were examined for vasculopathic changes and, in case of vasculitis, were further analyzed with electron microscopy and immunohistochemistry. Vasculopathy was observed in 76% of all COVID-19-related inflammatory skin lesions. Visual endothelial changes without manifest leukocytoclastic vasculitis were found in 60% of the COVID-19-related skin lesions, whereas leukocytoclastic vasculitis was diagnosed in 16%. In the cases of vasculitis, there were extensive spike protein depositions in microvascular endothelial cells that colocalized with the autophagosome proteins LC3B and LC3C. The autophagy protein complex LC3-associated endocytosis in microvascular endothelial cells seems to be an important pathogenic factor for severe acute respiratory syndrome coronavirus 2-related vasculitis in the skin. On ultrastructural morphology, the vasculitic process was dominated by intracellular vesicle formation and endothelial cell disruption. Direct presence of severe acute respiratory syndrome coronavirus 2 particles in the skin was not observed. Therefore, our results suggest that instead of direct viral infection, dermal vasculitic lesions in COVID-19 are caused by severe acute respiratory syndrome coronavirus 2 spike protein deposition followed by endothelial damage with activation of autophagy.

Nanoformulation-Based 1,2,3-Triazole Sulfonamides for Anti-Toxoplasma In Vitro Study.

Toxoplasma gondii is deemed a successful parasite worldwide with a wide range of hosts. Currently, a combination of pyrimethamine and sulfadiazine serves as the first-line treatment; however, these drugs have serious adverse effects. Therefore, it is imperative to focus on new therapies that produce the desired effect with the lowest possible dose. The designation and synthesis of sulfonamide-1,2,3-triazole hybrids (3a-c) were performed to create hybrid frameworks. The newly synthesized compounds were loaded on chitosan nanoparticles (CNPs) to form nanoformulations (3a.CNP, 3b.CNP, 3c.CNP) for further in vitro investigation as an anti-Toxoplasma treatment. The current study demonstrated that all examined compounds were active against T. gondii in vitro relative to the control drug, sulfadiazine. 3c.CNP showed the best impact against T. gondii with the lowest IC50 value of 3.64 µg/mL. Using light microscopy, it was found that Vero cells treated with the three nanoformulae showed remarkable morphological improvement, and tachyzoites were rarely seen in the treated cells. Moreover, scanning and transmission electron microscopic studies confirmed the efficacy of the prepared nanoformulae on the parasites. All of them caused parasite ultrastructural damage and altered morphology, suggesting a cytopathic effect and hence confirming their promising anti-Toxoplasma activity.

Observations on the ultrastructure of the oral appendages of Glenea cantor Fabricius (Coleoptera: Lamiinae).

Using scanning electron microscopy, we examined the gross and ultrastructure morphology of the mandibles, labial palpi, and mandibular palpi of adult male and female Glenea cantor beetles. The morphology of these parts, both in their gross and ultrastructure, varied significantly between males and females. The lengths of the mandible, labial palpi, and mandibular palpi were clearly noticeably longer in females than in men, which is mostly related to the oviposition mechanism. In terms of the ultrastructural morphology of the mandibles, labial palpi, and mandibular palpi, seven types of sensilla were found on these parts: sensilla twig basiconica (STB I, II, III, and IV), sensilla chaetica (Scheme IV and V), Böhm's bristles (Bb), sensilla placodea (SP), sensilla trichodea (ST II), sensilla plate (SP), sensilla coeloconica (SC), and sensilla campaniformia (SCa). Females have significantly more ST I and Scheme III on their mandibles than males. The mechanical sensors SCh and Bb, olfactory sensor ST II, taste sensor STB IV, and carbon dioxide sensor and temperature and humidity sensors SC, SP, and SCa were much more developed on the labial and maxillary palpi of females compared to those of males, which further clarified the significance of the oral appendages in oviposition behavior. These findings will advance information-based technique design and the creation of information-based pest control strategies by assisting in our understanding of the host preference and oviposition behavior of adult G. cantor. RESEARCH HIGHLIGHTS: The external morphology and distribution of the oral appendages of Glenea cantor between sexes has been researched through scanning electron microscopy (SEM) for the first time. Several significant differences between males and females have been found by analyzing the oral appendages of G. cantor with SEM. The differences in the structure of oral appendages of G. cantor between sexes reflect functional differences in reproductive behaviors.

Hypoosmosis alters hepatocyte mitochondrial morphology and induces selective release of carbamoyl phosphate synthetase 1.

Carbamoyl phosphate synthetase 1 (CPS1) is the most abundant hepatocyte mitochondrial matrix protein. Hypoosmotic stress increases CPS1 release in isolated mouse hepatocytes without cell death. We hypothesized that increased CPS1 release during hypoosmosis is selective and associates with altered mitochondrial morphology. Both ex vivo and in vivo models were assessed. Mouse hepatocytes and livers were challenged with isotonic or hypoosmotic (35 mosM) buffer. Mice were injected intraperitoneally with water (10% body weight) with or without an antidiuretic. Mitochondrial and cytosolic fractions were isolated using differential centrifugation, then analyzed by immunoblotting to assess subcellular redistribution of four mitochondrial proteins: CPS1, ornithine transcarbamylase (OTC), pyrroline-5-carboxylate reductase 1 (PYCR1), and cytochrome c. Mitochondrial morphology alterations were examined using electron microscopy. Hypoosmotic treatment of whole livers or hepatocytes led to preferential or increased mitochondrial release, respectively, of CPS1 as compared with two mitochondrial matrix proteins (OTC/PYCR1) and with the intermembrane space protein, cytochrome c. Mitochondrial apoptosis-induced channel opening using staurosporine in hepatocytes led to preferential CPS1 and cytochrome c release. The CPS1-selective changes were accompanied by dramatic alterations in ultrastructural mitochondrial morphology. In mice, hypoosmosis/hyponatremia led to increased liver vascular congestion and increased CPS1 in bile but not blood, coupled with mitochondrial structural alterations. In contrast, isotonic increase of intravascular volume led to a decrease in mitochondrial size with limited change in bile CPS1 compared with hypoosmotic conditions and absence of the hypoosmosis-associated histological alterations. Taken together, hepatocyte CPS1 is selectively released in response to hypoosmosis/hyponatremia and provides a unique biomarker of mitochondrial injury.NEW & NOTEWORTHY Exposure of isolated mouse livers, primary cultured hepatocytes, or mice to hypoosmosis/hyponatremia conditions induces significant mitochondrial shape alterations accompanied by preferential release of the mitochondrial matrix protein CPS1, a urea cycle enzyme. In contrast, the intermembrane space protein, cytochrome c, and two other matrix proteins, including the urea cycle enzyme ornithine transcarbamylase, remain preferentially retained in mitochondria. Therefore, hepatocyte CPS1 manifests unique mitochondrial stress response compartmentalization and is a sensitive sensor of mitochondrial hypoosmotic/hyponatremic injury.