Cytoplasmic Vacuoles Research Articles

Revealing the mechanism: the influence of Baicalin on M1/M2 and Th1/Th2 imbalances in mycoplasma gallisepticum infection

Mycoplasma gallisepticum (MG) is a pathogen that induces chronic respiratory illnesses in chickens, leading to tracheal and lung injury, and eliciting immune reactions that support sustained colonization. Baicalin, a compound found in scutellaria baicalensis, exhibits anti-inflammatory, antioxidant, and antibacterial properties. This study aimed to investigate the potential of baicalin in alleviating lung and cell damage caused by MG by restoring imbalances in M1/M2 and Th1/Th2 differentiation and to explore its underlying mechanism. In this research, a model for M1/M2 polarization induced by MG was initially developed. Specifically, infection with MG at a multiplicity of infection (MOI) of 400 for 6 h represented the M1 model, while infection for 10 h represented the M2 model. The polarization markers were subsequently validated using qRT-PCR, ELISA, and Western blot analysis. Baicalin disrupts the activation of M1 cells induced by MG and has the potential to restore the balance between M1 and M2 cells, thereby mitigating the inflammatory damage resulting from MG. Subsequent studies on MG-infected chickens detected imbalances in M1/M2 and Th1/Th2 differentiation in alveolar lavage fluid, as well as imbalances in macrophages and Th cells in the lung. The M1/Th1 model was exposed to MG for 5 d, while the M2/Th2 model was infected with MG for 7 d. The utilization of both light and electron transmission microscopes revealed that the administration of baicalin resulted in a reduction in the number of M1 cells, a decrease in cytoplasmic vacuoles, restoration of mitochondrial swelling and chromatin agglutination, as well as alleviation of alveolar rupture and inflammatory cell infiltration. Furthermore, baicalin restored MG-induced M1/M2 and Th1/Th2 imbalances and inhibited the phosphorylation of p38 and p65 proteins, thereby hindering the activation of the TLR4-p38 MAPK/NF-κB pathway. This study provides insights into the potential long-term effects of baicalin in MG infection and offers a theoretical basis for practical applications.

In situ fate of Chikungunya virus replication organelles.

Chikungunya virus (CHIKV) is a mosquito-borne pathogen responsible for an acute musculoskeletal disease in humans. Replication of the viral RNA genome occurs in specialized membranous replication organelles (ROs) or spherules, which contain the viral replication complex. Initially generated by RNA synthesis-associated plasma membrane deformation, alphavirus ROs are generally rapidly endocytosed to produce type I cytopathic vacuoles (CPV-I), from which nascent RNAs are extruded for cytoplasmic translation. By contrast, CHIKV ROs are poorly internalized, raising the question of their fate and functionality at the late stage of infection. Here, using in situ cryogenic-electron microscopy approaches, we investigate the outcome of CHIKV ROs and associated replication machinery in infected human cells. We evidence the late persistence of CHIKV ROs at the plasma membrane with a crowned protein complex at the spherule neck similar to the recently resolved replication complex. The unexpectedly heterogeneous and large diameter of these compartments suggests a continuous, dynamic growth of these organelles beyond the replication of a single RNA genome. Ultrastructural analysis of surrounding cytoplasmic regions supports that outgrown CHIKV ROs remain dynamically active in viral RNA synthesis and export to the cell cytosol for protein translation. Interestingly, rare ROs with a homogeneous diameter are also marginally internalized in CPV-I near honeycomb-like arrangements of unknown function, which are absent in uninfected controls, thereby suggesting a temporal regulation of this internalization. Altogether, this study sheds new light on the dynamic pattern of CHIKV ROs and associated viral replication at the interface with cell membranes in infected cells.IMPORTANCEThe Chikungunya virus (CHIKV) is a positive-stranded RNA virus that requires specialized membranous replication organelles (ROs) for its genome replication. Our knowledge of this viral cycle stage is still incomplete, notably regarding the fate and functional dynamics of CHIKV ROs in infected cells. Here, we show that CHIKV ROs are maintained at the plasma membrane beyond the first viral cycle, continuing to grow and be dynamically active both in viral RNA replication and in its export to the cell cytosol, where translation occurs in proximity to ROs. This contrasts with the homogeneous diameter of ROs during internalization in cytoplasmic vacuoles, which are often associated with honeycomb-like arrangements of unknown function, suggesting a regulated mechanism. This study sheds new light on the dynamics and fate of CHIKV ROs in human cells and, consequently, on our understanding of the Chikungunya viral cycle.

Astragaloside IV Alleviates Podocyte Injury in Diabetic Nephropathy through Regulating IRE-1α/NF-κ B/NLRP3 Pathway.

To investigate the effects of astragaloside IV (AS-IV) on podocyte injury of diabetic nephropathy (DN) and reveal its potential mechanism. In in vitro experiment, podocytes were divided into 4 groups, normal, high glucose (HG), inositol-requiring enzyme 1 (IRE-1) α activator (HG+thapsigargin 1 µmol/L), and IRE-1α inhibitor (HG+STF-083010, 20 µmol/L) groups. Additionally, podocytes were divided into 4 groups, including normal, HG, AS-IV (HG+AS-IV 20 µmol/L), and IRE-1α inhibitor (HG+STF-083010, 20 µmol/L) groups, respectively. After 24 h treatment, the morphology of podocytes and endoplasmic reticulum (ER) was observed by electron microscopy. The expressions of glucose-regulated protein 78 (GRP78) and IRE-1α were detected by cellular immunofluorescence. In in vivo experiment, DN rat model was established via a consecutive 3-day intraperitoneal streptozotocin (STZ) injections. A total of 40 rats were assigned into the normal, DN, AS-IV [AS-IV 40 mg/(kg·d)], and IRE-1α inhibitor [STF-083010, 10 mg/(kg·d)] groups (n=10), respectively. The general condition, 24-h urine volume, random blood glucose, urinary protein excretion rate (UAER), urea nitrogen (BUN), and serum creatinine (SCr) levels of rats were measured after 8 weeks of intervention. Pathological changes in the renal tissue were observed by hematoxylin and eosin (HE) staining. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were used to detect the expressions of GRP78, IRE-1α, nuclear factor kappa Bp65 (NF-κBp65), interleukin (IL)-1β, NLR family pyrin domain containing 3 (NLRP3), caspase-1, gasdermin D-N (GSDMD-N), and nephrin at the mRNA and protein levels in vivo and in vitro, respectively. Cytoplasmic vacuolation and ER swelling were observed in the HG and IRE-1α activator groups. Podocyte morphology and ER expansion were improved in AS-IV and IRE-1α inhibitor groups compared with HG group. Cellular immunofluorescence showed that compared with the normal group, the fluorescence intensity of GRP78 and IRE-1α in the HG and IRE-1α activator groups were significantly increased whereas decreased in AS-IV and IRE-1α inhibitor groups (P<0.05). Compared with the normal group, the mRNA and protein expressions of GRP78, IRE-1α, NF-κ Bp65, IL-1β, NLRP3, caspase-1 and GSDMD-N in the HG group was increased (P<0.05). Compared with HG group, the expression of above indices was decreased in the AS-IV and IRE-1α inhibitor groups, and the expression in the IRE-1α activator group was increased (P<0.05). The expression of nephrin was decreased in the HG group, and increased in AS-IV and IRE-1α inhibitor groups (P<0.05). The in vivo experiment results revealed that compared to the normal group, the levels of blood glucose, triglyceride, total cholesterol, BUN, blood creatinine and urinary protein in the DN group were higher (P<0.05). Compared with DN group, the above indices in AS-IV and IRE-1α inhibitor groups were decreased (P<0.05). HE staining revealed glomerular hypertrophy, mesangial widening and mesangial cell proliferation in the renal tissue of the DN group. Compared with the DN group, the above pathological changes in renal tissue of AS-IV and IRE-1α inhibitor groups were alleviated. Quantitative RT-PCR and Western blot results of GRP78, IRE-1α, NF-κ Bp65, IL-1β, NLRP3, caspase-1 and GSDMD-N were consistent with immunofluorescence analysis. AS-IV could reduce ERS and inflammation, improve podocyte pyroptosis, thus exerting a podocyte-protective effect in DN, through regulating IRE-1α/NF-κ B/NLRP3 signaling pathway.

Cytoplasmic vacuolization and ectopic formation of perineuronal nets are characteristic pathologies of cytomegalic neurons in tuberous sclerosis.

Cytomegalic neurons, characterized by increased size and a hyperactive mechanistic target of rapamycin complex 1 (mTORC1), are pathognomonic for tuberous sclerosis complex (TSC). To model these neurons, we recently generated a murine Tsc1 conditional knockout model in which Tsc1 deletion in late embryonic radial glia results in neuronal hypertrophy of a subset of isocortical pyramidal neurons. In the current study, we compared the cellular pathology of these cytomegalic neurons to those of the enlarged neurons in human cortical tubers. Neurons from the mice showed unique features, such as cytoplasmic vacuoles associated with Golgi complexes and the ectopic formation of perineuronal nets (PNNs), a feature of inhibitory neurons, rarely present in excitatory cortical neurons. The membranes of these vacuoles were enriched for the plasma membrane proteins CD44, KCC2, and Na+/K+ ATPase, suggesting deficits in Golgi membrane trafficking. These aberrant features in the mouse appeared only after the onset of seizures, probably due to the prolonged seizure activity in the context of constitutive mTORC1 activation. Similar PNNs and cytoplasmic vacuoles were present in the cytomegalic neurons of human cortical tubers. Our findings reveal novel pathological features of Golgi complexes and PNNs in the cytomegalic neurons in TSC.

Cytoplasmic Lipid Droplets Predict Worse Prognosis in Diffuse Large B-Cell Lymphoma: Next-Generation Sequencing Deciphering Lipogenic Genes.

Burkitt lymphoma is characterized by high cell turnover and numerous cytoplasmic vacuoles that are demonstrated to be lipid droplets (LDs) decorated by adipophilin. By contrast, cytoplasmic vacuoles are variably observed in diffuse large B-cell lymphoma (DLBCL) and less well characterized. In this study, we first validated in DLBCL that cytoplasmic vacuoles are indeed LDs by Oil-red-O stain, Bodipy fluorescent stain, and electron microscopy. Second, in a cohort of DLBCL patients (n=52) we showed that LDs in effusional lymphoma cells were associated with a poorer prognosis (P=0.029, log-rank test) and higher International Prognostic Index (IPI) score (94% vs. 66%, P=0.026) than those without. Moreover, using adipophilin as a surrogate marker for LDs, we found in another cohort of biopsy specimen (n=85) that expression of adipophilin by lymphoma cells predicted a poorer prognosis (P=0.007, log-rank test) and higher IPI score (63% vs. 30%, P=0.005). In addition, whole exome sequencing of effusional DLBCL cells showed LD-positive DLBCL shared genetic features with the MCD (MYD88 and CD79B mutations) subtype and highlighted OSBPL10 and CUBN as the most frequently mutated genes involved in lipogenesis. Whole transcriptome analysis by comparing effusional DLBCL cells with versus without LDs showed upregulation of EHHADH, SLC1A1, CD96, INPP4B, and RNF183 relevant for lymphoma lipogenesis and upregulation of epithelial-mesenchymal transition and KRAS signaling pathways. Higher expression of EHHADH and CD96 were validated in LD-positive clinical samples and LD-rich cell lines than LD-poor cells along with the known lipogenic gene, FASN. Our findings highlight the roles of LDs and adipophilin expression in DLBCL, suggest that these markers may predict prognosis and show that lipogenic genes may be potential therapeutic targets.

Imidacloprid‐associated toxicity in the midgut of Sarcophaga ruficornis: Exploring histopathological, ultrastructural and biochemical alterations

AbstractThis study investigates the impact of imidacloprid insecticide, on the flesh fly Sarcophaga ruficornis, (Diptera: Sarcophagidae), a medically and veterinary significant species associated with myiasis. Lethal (0.02%) and sub‐lethal (0.004%) doses were administered over 24 and 48 h, focusing on midgut histopathological, ultrastructural and biochemical alterations. Imidacloprid‐induced disruptions in physiological and metabolic functions, leading to adult fly mortality. Histological and cytological analyses revealed significant cellular changes, including cell degeneration, cytoplasmic vacuolization, chromatin condensation, irregular epithelial borders, disrupted peritrophic membrane and the release of cell components into the midgut lumen. Biochemical analyses demonstrated a decrease in total protein, carbohydrate and lipid contents, correlating with insecticide concentration and duration. Antioxidant enzymes, particularly glutathione S‐transferase (GST) and superoxide dismutase (SOD), resulted in a significant increase compared to the control, proportional to time and concentration. This research provides a comprehensive understanding of imidacloprid‐induced toxicity in the midgut of Sarcophaga ruficornis, emphasizing its potential for effective pest population management.

Amino-modified polystyrene nanoplastics induced multiple response of Artemia hemocytes

Polystyrene polymers cause severe toxicity to aquatic animals. However, the process and mechanisms of innate immunity of invertebrates living at the bottom of the food chain to these pollutants remain unclear. In this study, the blood system responses of zooplankton Artemia were assessed through in vivo and in vitro exposure to amino-modified polystyrene nanoplastics (PS-NH2 NPs). The results indicated that the LC50 values of PS-NH2 NPs were 1.09 μg·mL−1 over 48 h and 0.42 μg·mL−1 over 7 d. Based on the five hemocyte subpopulations identified in Artemia, in vitro exposure assays revealed that phagocytosis was performed by plasmocytes and granulocytes with phagocytic rate of 22.64 %. TEM analysis further showed that PS-NH2 NPs caused cytoplasm vacuolization, swollen mitochondria, and lipid processing disorder. Gene expression pattern results demonstrated that Spatzle, Tollip, Hsp70, Hsp90, Casp8, API5and Pxn were significantly upregulated upon acute and chronic exposure (p < 0.05), while chronic exposure could induce significantly upregulation of ProPO (p < 0.05). Moreover, PS-NH2 NPs exposure remarkably varied the hemolymph microbiota and hemogram, particularly by increasing the proportion of adipohemocytes and phagocytes (p < 0.05). Our findings suggest that PS-NH2 NPs induce different responses in Artemia hemocyte, as primarily reflected by phagocytic processes, expression of immune and apoptosis relating genes, cell fates, hemogram and hemolymph microbiota variations. These findings support the possibility of using Artemia hemocytes as bioindicator to estimate nanoplastics pollution, thus contributing to hematological toxicity research in response to nanoplastics.

Protective Effects of Stem Cells and Vitamin D3 Against Hepatic Ischemia/Reperfusion Injury in Rats

Abstract Background Liver transplantation is one of the most efficient life-saving treatments for various end-stage hepatic diseases. However, hepatic ischemic reperfusion injury (IRI) remains a major problem in liver surgeries morbidities and mortalities. Stem cells (SCs) have been recently received increasing attention for cell-based therapy, especially in regenerative medicine, including liver diseases. Aim to investigate the effect of vitamin D and Bone Marrow Mesenchymal Stem Cells (BM- MSCs) solely or combined on hepatic IR injury. Methods Sixty adult male Wister rats were allocated into 5 groups; Sham, IR: underwent hepatic IR, vit. D-IR: received vit. D in a dose of 500 IU/Kg/day, by gavage, 5 days/week for 2 weeks, then subjected to hepatic IR, Stem cell-IR: received single IV injection of 1 × 106 BM-MSCs prior to hepatic IR and Combined-IR: received both vit. D and BM-MSCs prior to hepatic IR. All rats were subjected to estimation of Liver index (LI), assessment of serum levels of alanine, aspartate transaminases (ALT, AST) and vitamin D, hepatic tissue levels of Malondialdehyde (H.MDA), Superoxide dismutase (H.SOD), Interleukin-2, 10 (H.IL-2, H.IL-10), Caspase-3 (H.Casp-3), H.LC3B and hepatocyte growth factor (H.HGF), as well as histopathological liver examination and scoring. Results Hepatic IR resulted in significant elevation of ALT, AST and H.MDA, H.IL-2, H. Casp- 3 as well as significant reduction in LI, vit. D, H.SOD, H.IL-10, H.LC3B and H.HGF with evident hepatic sinusoidal congestion, cytoplasmic vacuolation and cellular necrosis. Treatment either solely or in combination mitigate these changes with the combined treatment showed the best results compared to the individual use. Conclusion In vivo coadministration of vitamin D and BM-MSCs prior to hepatic IR attenuated hepatocellular damage through preventing oxidative stress and inflammatory response, augmenting autophagic, regenerative capacity and anti-apoptotic effect.

P-362 Cellular senescence in mouse ovaries as an explanatory mechanism of cyclophosphamide detrimental effect to female fertility

Abstract Study question Is cellular senescence a causative mechanism of cyclophosphamide’s detrimental effect to female fertility? Summary answer Cyclophosphamide induced up-regulation of genes related to cellular senescence and associated pathways, oocyte organelles alterations and increased beta-galactosidase (b-Gal) positive cells in mouse ovaries. What is known already Cyclophosphamide (CPA) is frequently used for treatment of cancer and chronic benign diseases; however, secondary effects include infertility and POI. The mechanisms of CPA-induced POI have not been fully elucidated. Research in possible causative mechanisms include apoptosis and over-activation with follicle burnout, however, new hypothesis including affecting cellular senescence has been proposed. Cellular senescence is a complex cellular reaction to stress and the cells enter an irreversible proliferative arrest state. It is associated with alterations in cell secretory profile (secretion of pro-inflammatory factors) and morphology (vacuolization and granularity in cytoplasm, abnormal organelles). Study design, size, duration Controlled experimental study. Intact ovaries from B6CBA/F1 4-day-old mice were randomly assigned to 4-hydroperoxycyclophosphamide (n = 22) or control group (n = 22). Five ovaries per group were collected at 8, 12, 24, 36 h and processed for RNA sequencing. Two ovaries per group were collected at 24 h for transmission electron microscopy analysis. B6CBA/F1 12-day-old mice were either intraperitoneally injected with 100 mg/kg CPA (n = 2) or no treatment (n = 2). Ovaries were collected for histological analysis 2 weeks later. Participants/materials, setting, methods Ovaries from 4-day-old mice were cultured on Millicell cell culture inserts floating on 0.25 mL culture medium in a 24-well plate, maximum 5 ovaries per insert. Freshly prepared 4-hydroperoxycyclophosphamide solution was added to the wells of CPA group (final concentration is 5 µM). 12-day-old mice were intraperitoneally injected with 100 mg/kg CPA or no treatment, 2 weeks later, ovaries were collected and fixed, then processed for b-Gal staining. Main results and the role of chance Transcriptomic analysis of 4-day-old ovaries cultured with 4-hydroperoxicyclophosphamide (5 µM) presenting up-regulation of genes related to cellular senescence and its associated signaling pathways, e.g. TNF signaling pathway, P53 signaling pathway, cell cycle, cytokine-cytokine receptor interaction. Literature reported cellular senescence related genes Cdkn1a, Hdac1, E2f3, Ccne1, Jun, Pcna, Smad3, Mapk1, Smad1, Cdkn2cc, Ccne2, Rbl1, Tp53, Myc, Cdk2, Cdk1 were found up-regulated by CPA treatment at 36 h. CPA treatment also caused organelles alterations in ovarian follicles revealed by transmission electron microscopy, e.g. endoplasmic reticulum became elongated and attached to mitochondria and forming a necklace-like structure, vacuolization could be observed in some oocytes’ cytoplasm, especially in some growing follicles. And these changes were not observed in the control group. Young (12-day-old) mice that received CPA (100 mg/kg) showed increased trend of staining of cellular senescence markers including b-Gal two weeks after the treatment comparing to control ovaries, suggesting an involvement of cellular senescence in CPA induced ovarian damage. Limitations, reasons for caution We used an experimental model and it is not known if similar results could be found in human ovarian tissue. Wider implications of the findings Our results using in vitro and in vivo models suggest that cellular senescence may be induced by CPA at transcriptomic, cellular ultrastructural and histological levels. This might partially explain the occurrence of infertility following treatment with alkylating drugs since some ovarian cells are arrested and influence follicle development. Trial registration number Not Applicable

P-164 The impact of oocyte morphological abnormalities on fertilization rate and blastocyst quality

Abstract Study question Do distinct morphological abnormalities in oocytes have an adverse effect on both fertilization rates and the quality of resulting blastocyst? Summary answer While most oocyte morphological abnormalities did not significantly affect outcomes, dark cytoplasmic color correlated with lower fertilization rates and the absence of good quality blastocysts What is known already Variability exists in the fertilization and developmental potential of mature (metaphase II - MII) human oocytes. A high-quality MII oocyte is characterized by a spherical shape, homogeneous cytoplasm, smooth appearance, a uniform zona pellucida (ZP), a normal-sized perivitelline space (PVS), and a single unfragmented polar body (PB). Dysmorphisms, such as variations in cytoplasmic features and anomalies in PVS size and PB regularity may occur. There are conflicting reports on the relationship between oocyte morphology and IVF outcomes. A recent meta-analysis highlighted associations between poor fertilization potential and specific dysmorphisms, including large/irregular PB, a large PVS, refractile bodies, and vacuoles. Study design, size, duration This study analyzed 367 MII oocytes obtained from 40 young patients. The fertilization rate (FR) and blastocyst quality (BQ) were compared among 132 dysmorphic and 235 normal MII oocytes (from October 2023 to January 2024). Dysmorphic oocytes were categorized as having a non-spherical shape, large PVS, PVS debris, abnormal ZP, fragmented PB, excess cytoplasmic granules, vacuoles, and dark cytoplasmic color. Participants/materials, setting, methods All MII oocyte denuded were examined under the microscope at x-400 magnification. Dysmorphic and normal oocytes were separated, injected, and cultured individually until day 5 under conditions of 37oC and 6% CO2. Fertilization was assessed 14 to 18 hours post-injection, and subsequent embryonic development was evaluated on day 2 and day 5. Blastocysts graded 4AA to 4BC are considered good quality, while those graded 4CC to 3CC are classified as fair quality. Main results and the role of chance The mean age of patients was 24.14 ± 3.77. In total, 36% of all oocytes exhibit at least one dysmorphism (n = 132). From the overall dataset analyzed, there was no significant difference (p &amp;gt; 0.9999) in FR between the normal 77.87% (183/235) and dysmorphic group of oocytes 78.03 (103/132). The odd ratio and 95% confidence interval are 0.9909 and 0.5957 to 1.633 respectively. However, this was not the same for oocytes with dark color cytoplasm. The presence of this dysmorphism significantly lowered FR (p = 0.0176) with an odd ratio of 5.544 (95% CI: 1.419 to 21.67). Similarly, the result showed no statistically significant difference (p = 0.1073) in the likelihood of obtaining a fair quality blastocyst between the normal oocytes 23.40% (55/235) and dysmorphic group of oocytes 15.79 (21/133). This was also the same for the likelihood of obtaining good quality blastocyst. There was no statistically significant difference (p = 0.1536) between the normal group 20.00% (47/235) and the dysmorphic group 26.52 (35/132). Oocyte with dark color cytoplasm gave no good quality blastocyst. Limitations, reasons for caution The exclusive use of oocytes from young patients may limit the generalizability of findings to a broader population of IVF patients. Also, the sample size may limit the study’s ability to detect subtle associations. A reason for caution when interpreting this study is its focus on morphological assessments alone. Wider implications of the findings The knowledge from our findings may contribute to optimizing oocyte selection strategies for intracytoplasmic sperm injection. Also, the knowledge could potentially be integrated into AI modules for oocyte morphological assessment. However, more studies would be required to develop a robust and scalable system Trial registration number Not applicable

The Possible Protective Role of Resveratrol against Zinc Oxide Nanoparticles Induced Hepatic Tissue Injury in Adult Albino Rat

Abstract Background Zinc oxide nanoparticles (ZnO NPs) are widely used in industries and medications. This may elevate the probability of their existence in the environment, which may have harmful side effects on different body organs, especially on the liver. So it became mandatory to search for natural hepatoprotective elements such as Resveratrol. Aim of the Work The aim of the present work was to determine the effect of zinc oxide nanoparticles on the histological structure and biochemical markers of the liver of adult male albino rats and to evaluate the possible protective role of Resveratrol. Material and Methods 40 adult male albino rats aged 3-6 months, were divided randomly into four groups; Group I (Control group), which consisted of 10 male albino rats with free access to food (rat chew) and water; Group II (ZnO NPs group), which included 10 male albino rats, which received a dose of 50 mg/kg body weight ZnO NPs through oral gavage daily for 4 weeks; Group III (Resveratrol group) was composed of 10 male albino rats which received a dose of 20 mg/kg body weight Resveratrol through oral gavage daily for 4 weeks, and Group IV (ZnO NPs + Resveratrol group) included 10 male albino rats which were co-administered with a combination of ZnO NPs at a dose of 50mg/kg body weight and Resveratrol at a dose of 20 mg/kg body weight dose through oral gavage daily for 4 weeks. At the end of the experiment, blood samples were collected for serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP). All the animals were sacrificed, and the liver was extracted and processed into paraffin blocks for light microscopic examination. A morphometric study and a statistical analysis were done. Results The present work demonstrated that liver sections obtained from adult male albino rats that received ZnO NPs showed disorganized hepatic architecture, severe affection of the hepatocytes, which had small darkly stained nuclei, and deep acidophilic cytoplasm. The hepatocytes sometimes showed extensive cytoplasmic vacuolations. Most hepatocytes had small apoptotic nuclei and were surrounded by clear halos. While some hepatocytes lost their cell boundaries and coalesced together. Central veins were dilated, distorted, and congested with areas of epithelial lining detachment. Most of the blood sinusoids appeared congested with marked inflammatory cell infiltration. The portal tracts showed vascular congestion, dilatation, and proliferation of the bile ducts. There was a marked increase in the collagen deposition around the portal tracts, central veins, and in-between hepatocytes. Marked depletion of glycogen content with sporadic positive PAS reactions within the cytoplasm of the hepatocytes was also noticed. Hepatocyte apoptosis was evident by a significant increase in caspase-3 expression compared to the control group. On the other hand, ZnO NP +Resveratrol group showed the restoration of normal hepatic lobules with few areas of subcapsular haemorrhage. Most hepatocytes appeared polygonal in shape, with acidophilic cytoplasm and central rounded vesicular nuclei. While few hepatocytes appeared degenerated with dark stained pyknotic nuclei and cytoplasm showed hydropic degeneration. Most of the blood sinusoids returned to their normal sizes with flattened endothelial cells. However, sometimes they were congested, dilated, and contained Kupffer cells. Some central veins appeared congested. The portal tract also returned to its normal size with a marked decrease in inflammatory cell infiltration and showed apparent minimal collagen fiber deposition. Another finding in the present work was that Resveratrol restored the glycogen content of the hepatocytes which were depleted in ZnO NPs group. This was marked by a strong PAS positive reaction in the cytoplasm of most hepatocytes, with few hepatocytes with weak PAS reaction. Resveratrol co-treatment with ZnO NPs significantly reduced the degeneration and necrosis of hepatocytes. This was evident in our study by the apparent negative Caspase-3 reaction in the cytoplasm of most hepatocytes. Clear deterioration of liver function tests was observed in ZnO NPs group compared with the control, with marked improvement in the group treated with ZnO NPs and Resveratrol together. Conclusion The present study demonstrated the degenerative and hepatotoxic effects of ZnO NPs on the histological structure and function of the liver and the hepatoprotective role of Resveratrol against these effects.

Effect of Consumption of Caffeinated Energy Drinks on the Myocardium of the Adult Male Albino Rat (A Histological and Morphometric Study)

Abstract Background Energy drinks are non-alcoholic beverages containing mainly caffeine. Chronic consumption of energy drinks may be associated with increased blood pressure, and cardiac work load, as well as, increased heart rate. Energy drinks consumption has been related to myocardial infarction in healthy young adult. Aim of the Work The aim of the present work was to study the harmful effect of energy drinks histologically and morphometrically, on the myocardium of the adult male albino rats. Material and Methods 20 Adult male albino rats aging 4-6 moths were divided randomly into two groups; Group I (Control group) which consisted of 10 male albino rats with free access to food (rat chew) and water and Group II (Experimental group): 10 male albino rats which received 3.75 ml/kg body weight red bull through oral gavage daily for 4 weeks with free access to food (rat chew) and water. At the end of experiment, rats werescarificed, and cardiac muscle was extracted and processed into paraffin blocks for light microscopic examination. Morphometric study and statistical analysis were done. Results The present work demonstrated that Red Bull induced several histopathological changes of the cardiac muscle. The results showed the cardiac muscle of rats with irregularities in the arrangement of fibers and wide spaces in between. Some cardiomyocytes showed areas of pale stained sarcoplasm, partial loss of the cross striations and ill-defined intercalated discs. Localized areas of hyaline degeneration and areas of tissue disruption were also noticed with less frequently localized coalesced cytoplasmic vacuoles. The nuclei of the cardiomyocytes appeared small, darkly stained, and pyknotic. Others were flattened and deeply stained. Congested, and thick walled blood vessels were also observed, with mononuclear cellular infiltrations. In Masson’s Trichrome sections, relative increase in collagen deposition between the cardiac muscle fibers was observed which was associated with focal fibrous degeneration. Examination of the semithin sections showed partial disruption of the cardiomyocytes. The nuclei of the cardiomyocytes were small, darkly stained with perinuclear degeneration. Some nuclei were eccentric, or with an irregular outline, and peripheral chromatin condensation. The characteristic finding was the presence of longitudinal wavy cardiac muscle fibers with relatively wide spaces in between and localized areas of partially lost cross striations and intercalated discs. Dilated, thick walled blood vessels was noticed between the bundles of cardiac muscle. Mononuclear inflammatory cellular infiltrations were also detected. Conclusion The present study demonstrated the degenerative and cardiotoxic effects of Red Bull as an example of energy drinks on the histological structure of the cardiac muscle.

Clinical Characteristics and Diagnosis of Nonaccelerating MDS/MPN-U Patient with Granulocyte Dysplasia.

The goal was to improve the clinical cognition of nonaccelerating myelodysplastic/myeloproliferative neoplasms-unclassifiable (MDS/MPN-U) and avoid misdiagnosis or delayed diagnosis. The clinical manifestations, laboratory indicators, histopathology, and therapeutic effects of a patient with nonaccelerating MDS/MPN-U were analyzed and the relevant literature was reviewed. Blood routine: white blood cell 98.48 x 109/L, red blood cell 3.20 x 1012/L, basophils 0.42 x 109/L, eosinophils 1.31 x 109/L, hemoglobin 112 g/L, and platelet 113 x 109/L. Blood smears showed granulocytosis and cells at various stages, polylobular granulocytes also can be seen. Bone marrow images show granulocytosis and dysplastic neutrophils, such as binuclear granulocyte, cyclic nuclear granulocyte, nuclear punch, cytoplasm vacuoles, polylobular granulocytes and so on. Bone marrow biopsy: Bone marrow proliferation tumor, combined with cell morphology and molecular biochemistry is recommended. Gene test showed Jak-2 positive, BCR/ABL and MPL negative. Chromosome examination indicated the presence of 46, XY, add (2)(p25), del (12) (p11.2p13)[16]/46, XY. MDS/MPN-U with granulocytosis and dysplastic neutrophils is rare, mostly in the elderly, and the diagnosis should be made except for other myeloid tumors. Currently, there is no uniform treatment guideline or expert consensus. The treatment options are limited and need to be further confirmed by more studies. MDS/ MPN-U with granulocytosis and dysplastic neutrophils has adverse prognostic factors such as advanced age, increase of bone marrow original cells and related gene mutations. Whether the adverse prognosis is related to specific gene mutations and cytogenetic variation remains to be clarified by more research data.

The potential therapeutic effects of exosomes derived from bone marrow mesenchymal stem cells on ileum injury of a rat sepsis model (histological and immunohistochemical study)

ABSTRACT Sepsis denotes a serious high mortality concern. The study was designed to evaluate the effect of mesenchymal stem cell exosomes (MSC-exosomes) on the evolution of the animal model of sepsis. In this study, 36 rats were distributed into three groups, (I) controls, (II) LPS-treated, and (III) LPS+MSC-EVs. Sepsis was simulated by administering E. coli-LPS to the laboratory animals. Group III was given MSC-exosomes four hours after the LPS injection. Forty-eight hours later rats were sacrificed. Ileum samples were excised, and processed for the histological assessment, immunohistochemical identification of CD44, and inducible nitric oxide synthase (iNOS). Ileum homogenate was used to estimate tumor necrosis factor α (TNF α) besides Cyclooxygenase-2 (COX 2). PCR was used for the detection of interleukin 1α (IL‑1α), and interleukin 17 (IL‑17). Statistical and morphometrical analysis was done. The LPS-treated group showed increased TNF-α, IL‑1α, IL‑17, and decreased COX 2. LPS administration led to cytoplasmic vacuolization of enterocytes, an increase in the vasculature, and cellular infiltrations invaded the lamina propria. There was a significant rise in goblet cells and the proportion of collagen fibers. Ultrastructurally, the enterocytes displayed nuclear irregularity, rough endoplasmic reticulum (rER) dilatation, and increased mitochondria number. Sepsis induces a significant increase in iNOS and a decrease in CD44 immune expressions. LPS+MSC-EVs group restored normal ileum structure and revealed a significant elevation in CD44 and a reduction in iNOS immunoreactions. LPS-sepsis induced an obvious ileum inflammatory deterioration ameliorated by MSC-exosomes, mostly through their antioxidant, anti-inflammatory, and anti-apoptotic properties.

Incidence and pathological effects during encysted progenetic Euclinostomum heterostomum metacercariae infection in the freshwater fish spotted snakehead (Channa punctatus)

Aim: To assess the incidence and pathological effects during encysted progenetic Euclinostomum heterostomum metacercariae infestation in the freshwater fish spotted snakehead (Channa punctatus). Methodology: Samples of Channa punctatus were collected from Tilaiya Reservoir, Koderma, Jharkhand, India, during pre-monsoon, monsoon, and post-monsoon seasons. The encysted metacercariae trematodes were isolated from Channa punctatus. Characterization was performed based on the morphology and sequencing analysis of a partial 18S rRNA gene. Further, the kidney and liver samples from infected fish were subjected to histopathological studies. Prevalence, abundance, and Intensity of infection were further determined. Results: Morphological identification suggests that the encysted metacercariae belonged to Digenean trematodes of genera Euclinostomum, and amplification of partial sequence of 18S rRNA, and phylogenetic analysis revealed the isolated metacercariae were Euclinostomum heterostomum. The histopathological study of the kidney showed hepatocyte degeneration, cytoplasmic vacuolation, and nuclear transformations. Liver fibrosis was observed in Channa punctatus. The prevalence during pre-monsoon, monsoon, and post-monsoon season ranged from 66.1% to 81.1%, abundance from 0.74 to 0.92, and intensity ranged from 1 to 1.14 using the Margolis formula. Interpretation: The study highlights the infestation of Euclinostomum heterostomum in Channa punctatus during the pre- monsoon climate appears favorable for the abundance of parasites. Hence, a proper inspection before consumption is necessary. Key words: Euclinostomum heterostomum, Infestation, Metacercariae, Parasites, Progenetic infection

Occurrence of Microplastics in the Tissues of Nile Tilapia (Orechromis niloticus) from Zobe Dam, Katsina State, Nigeria

A study of the detection of microplastics on Oreochromis niloticus in Zobe Reservoir, Katsina State, Nigeria was carried out. Three fish sampling sites were used; the upstream (Tsawatsawa), midstream (Makera), and downstream (Garhi) were selected based on variation of anthropogenic activities. A total number of one hundred and forty-four (144) fish samples with a total weight (MTW) and total length (MTL) varied between 134.1 ±1.50 to 154.2± 3.41g and 17.5 ± 0.10– 22.1± 0.54 cm respectively, were collected and analysed for the presence of microplastics using standard protocols. Fourier Transform Infrared Spectrophotometer (FTIR) was used for the chemical characterization of microplastic composition examined in the fish species. The examination of microplastic occurrence in different sampled fish gills, liver, and kidney revealed the presence of Polystyrene, Polypropylene, Polyethylene terephthalate, Nylon, Polyester, and Polyvinyl chloride. Fish from all sampling sites revealed varying histopathological alterations ranging from lifting of epithelium thickening, distortion of the primary lamellae, and degeneration of secondary lamellae, blood congestion, and massive lamellae degeneration in the gill tissues. Liver tissues had hepatocyte hypertrophy, sinusoidal dilatation and cytoplasmic vacuolation, presence of lipid granules, cellular infiltration, cytoplasmic vacuolation, and haemorrhage. The kidney tissues were presented with Bowman’s capsules hyperplasia, tubular elongation, renal tubular epithelium shrinkage, hyperplasia, and cytoplasmic vacuolization of renal cells. The severity of the damage varies from site to site indicating that the water body is experiencing pollution at varying locations. Our findings provide important insights into the prevalence and distribution of microplastics in this particular environment.

Therapeutic potential of silver nanoparticles from Helianthemum lippii extract for mitigating cadmium-induced hepatotoxicity: liver function parameters, oxidative stress, and histopathology in wistar rats.

Introduction: This study explores the therapeutic potential of silver nanoparticles (Ag NPs) synthesized using a Helianthemum lippii extract in mitigating cadmium-induced hepatotoxicity in Wistar rats. Given the increasing environmental and health concerns associated with cadmium exposure, novel and eco-friendly therapeutic strategies are essential. Methods: Ag NPs were characterized using X-ray diffraction, UV-Vis spectrometry, and energy-dispersive X-ray spectroscopy with scanning electron microscopy, confirming their formation with a cubic crystal structure and particle sizes ranging from 4.81 to 12.84nm. A sub-acute toxicity study of Ag NPs (2mg/kg and 10mg/kg) was conducted, showing no significant difference compared to untreated control rats (n = 3 animals/group). Subsequently, adult Wistar rats (n = 5/group) were divided into a control group and three experimental groups: Ag NPs alone, exposure to 50mg/kg CdCl2 in drinking water for 35days, and CdCl2 exposure followed by 0.1mg/kg/day Ag NPs intraperitoneally for 15days. Results: In the CdCl2-exposed group, there was a significant decrease in body weight and increases in alanine and aspartate transaminase levels (p < 0.05 vs. control), indicating hepatotoxicity. Additionally, antioxidant defenses were decreased, and malondialdehyde levels were elevated. Liver histology revealed portal fibrosis, inflammation, necrosis, sinusoid and hepatic vein dilation, and cytoplasmic vacuolations. Treatment with Ag NPs post-CdCl2 exposure mitigated several adverse effects on liver function and architecture and improved body weight. Discussion: This study demonstrates the efficacy of Ag NPs synthesized via a green method in reducing cadmium-induced liver damage. These findings support the potential of Ag NPs in therapeutic applications and highlight the importance of sustainable and eco-friendly nanoparticle synthesis methods. By addressing both toxicity concerns and therapeutic efficacy, this research aligns with the growing emphasis on environmentally conscious practices in scientific research and healthcare.

Revised Diagnosis From Histiocytic Neoplasm to Optic Chiasm Glioblastoma After Genetic Analysis.

A 46-year-old man presented with left eye blurring. Automated visual field testing showed an incongruous right hemianopia, with sparing of the lower temporal quadrant in the right eye. MRI revealed foci of gadolinium enhancement in the optic chiasm and optic tracts. Serologic testing (including myelin oligodendrocyte glycoprotein and neuromyelitis optica antibodies) and cerebrospinal fluid analysis were negative. Whole-body PET/CT scan found no malignancy. Biopsy of the optic chiasm revealed a moderately cellular neoplasm composed of atypical, discohesive cells with enlarged nuclei, prominent eosinophilic nucleoli, and abundant vacuolated cytoplasm. Immunohistochemical stains for CD68 and S100 were positive, whereas those for GFAP, OLIG2, SOX10, and multiple others were negative, supporting a diagnosis of histiocytic neoplasm. Five weeks later, results became available from next-generation sequencing targeting the coding regions of hundreds of malignancy-associated genes and select introns. Alterations associated with histiocytic neoplasms (i.e. BRAF and MAP2K1 mutations) were absent. However, there was a nonsense mutation in the PTEN gene, a hotspot mutation in the TERT gene promotor, and focal amplifications of the CDK4 and MDM2 genes. Additionally, there was chromosome 6q loss, 7 gain, and 10q loss. Based on these findings, the diagnosis was revised to glioblastoma, IDH-wildtype, CNS WHO grade 4. The patient began treatment with temozolomide while continuing radiation therapy. This case illustrates how next-generation sequencing can at times provide more accurate diagnostic information than standard tissue histopathology.

Biochanin A Has Protective and Therapeutic Effects against Li2CO3-induced Nephrotoxicity in Rats Model

BACKGROUND: Long-term usage of lithium carbonate (Li2CO3) as treatment of bipolar disorder might cause nephrotoxicity. Li2CO3-induced nephrotoxicity usually treated by renal dialysis, which is quite expensive. Usage of natural products that possess anti-inflammatory and antioxidant properties as biochanin A (BCA) may be useful against Li2CO3 nephrotoxicity. Therefore, this study was conducted to evaluate Li2CO3 nephrotoxic effects and BCA therapeutic and protective effects against it.METHODS: Twenty-eight rats divided into 4 groups: negative control (NC), Li2CO3, Li2CO3 + BCA therapeutic, and BCA + Li2CO3 protective group. Complete blood picture, serum kidney function tests (urea, creatinine, and cystatin C) and kidney tissue homogenate oxidative stress index as malonaldehyde (MDA), superoxide dismutase (SOD), glutathione S-transferase (GST) and catalase (CAT) were measured using enzyme linked immunosorbent assay (ELISA) methods. While, kidney histopathological changes were evaluated under light microscope.RESULTS: Li2CO3 administration elicited significant changes in hematological parameters as hypochromic anemia, decreased platelets and leukocytosis. Serum urea, creatinine and cystatin C were elevated. Renal homogenate analysis revealed significant elevation of MDA and decline in GST, CAT, SOD levels. Kidney histopathology analysis revealed hemorrhage between renal tubules, glomerular capillaries congestion, Bowman’s space dilatation, structural alterations in renal tubules cells include loss of brush border, deeply stained nuclei and elevated in size and number of cytoplasmic vacuoles. Therapeutic and protective groups of BCA showed amelioration of hematological alteration, kidney functions, oxidative stress markers and restoration of kidney tissues.CONCLUSION: BCA has both protective and therapeutic effects against Li2CO3-induced nephrotoxicity possibly through the reduction of oxidative stress.KEYWORDS: biochanin A, hematological changes, kidney, lithium carbonate salt, oxidative stress, protective effects, therapeutic effects

Development and Holocrine Secretion of Resin Ducts in Kielmeyera appariciana (Calophyllaceae).

The modes of formation and release of secretion are complex processes that occur in secretory ducts and their description has great divergence in some species. The use of modern techniques to detect hydrolytic enzymes, cytoskeleton arrangement and indicators of programmed cell death may help clarify the processes involved during the ontogeny of that gland. The goal of our study was to analyze subcellular changes during schizogenous formation and secretion production and release into the lumen in resin ducts of Kielmeyera appariciana. Our results demonstrate the participation of pectinase through the loosening of the central cells of the rosette, which subsequently split from each other through polarized growth mediated by a rearrangement of the microtubules. The resin is mainly synthesized in plastids and endoplasmic reticulum and is observed inside vesicles and small vacuoles. The secretion release is holocrine and occurs through programmed cell death related to the release of reactive oxygen species, causing cytoplasm darkening, chromatin condensation, vacuole rupture and plastid and mitochondria degeneration. Cellulase activity was identified prior to the rupture of the cell wall, causing the release of secretion into the lumen of the duct. The participation of the cytoskeleton was observed for the first time during schizogeny of ducts as well as programmed cell death as part of the process of the release of holocrine secretion. This type of secretion release may be a key innovation in Kielmeyera since it has not been observed in ducts of any other plant thus far.