Apoptotic Bodies Research Articles

Characterization of esophageal biopsies from stem cell transplant patients with and without esophageal graft-versus-host disease.

Graft-versus-host disease (GVHD) is a major complication of hematopoietic stem cell transplantation (HSCT). Histologic diagnostic criteria and several grading systems have been described for colonic GVHD; however, for esophageal GVHD (eGVHD) limited reports exist to date. In this study, a total of 130 HSCT patients with esophageal biopsies were included, with a median age of 44 years (2-77) and a male predominance (54.6%). Of these, 82 (63%) had a clinical diagnosis of eGVHD. Cases were divided into two groups: those without apoptotic bodies, dyskeratotic cells or ulceration (group 1, no histologic evidence of eGVHD) (42%) and those with at least one of those features (group 2) (58%). Group 2 cases were associated with extra gastrointestinal tract GVHD (p=0.024), a clinical diagnosis of eGVHD (p=0.001), older age (p<0.001), stem cells derived from peripheral blood (p<0.001), higher number of intraepithelial lymphocytes (p=0.002), presence of acute inflammation (p<0.001) and basal cell hyperplasia (p=0.016). Apoptotic bodies were seen in 65 (89%), dyskeratotic cells in 27 (37%) and an ulcer in 28 (37%) of the group 2 cases. The sensitivity (Sn), specificity (Sp) and accuracy (acc) of the group 2 cases for a clinical diagnosis of eGVHD was 68.3%, 60.4% and 65.4%, respectively. Apoptotic bodies (p=0.012) and dyskeratotic cells (p<0.001) but not ulceration (p=0.881), were associated with a clinical diagnosis of eGVHD. The Sn, Sp and acc for apoptotic bodies, dyskeratotic cells and ulcer were 59.3%, 63.8% and 60.9%, 30.9%, 95.7% and 54.7%, 21.9%, 79.2% and 43.1%, respectively. Cases with only apoptotic bodies or ulceration were considered as possible GVHD, and those with dyskeratotic cells as likely GVHD, which were associated with GVHD specific survival (p=0.030). This study provides a comprehensive characterization of the esophageal histologic findings in HSCT patients. Further studies are needed to corroborate these findings in other patient populations.

Unlocking the Potential of Extracellular Vesicles in Cardiovascular Disease.

Extracellular vesicles (EVs) are micro-nanoscale biological particles encapsulated by phospholipid bilayers, which regulate cell migration, angiogenesis and tumour cell growth by transmitting various biomolecules such as nucleic acids and proteins. EVs are composed of exosomes, microparticles and apoptotic bodies. Its benefits pass through biofilms and are not degraded by various enzymes, so it can be used as a biomarker in potential diseases and has attracted much attention from researchers. Current studies have found that EVs are involved in the development of various cardiovascular diseases (CVD), such as heart failure and myocardial ischemia-reperfusion injury. In addition, stem cell-derived EVs play an important role in the diagnosis and treatment of a variety of CVD. In this review, we present the biological features of EVs, the role of EVs in various CVD, and the challenges they encounter in the treatment of CVD.

Maternal milk cell components are uptaken by infant liver macrophages via extracellular vesicle mediated transport.

Milk is a multifaceted biofluid that is essential for infant nutrition and development, yet its cellular and bioactive components, particularly maternal milk cells, remain understudied. Early research on milk cells indicated that they cross the infant's intestinal barrier and accumulate within systemic organs. However, due to the absence of modern analytical techniques, these studies were limited in scope and mechanistic analysis. To overcome this knowledge gap, we have investigated the transintestinal transport of milk cells and components in pups over a 21-day period. Studies employed a mT/mG foster nursing model in which milk cells express a membrane-bound fluorophore, tdTomato. Using flow cytometry, we tracked the transport of milk cell-derived components across local and systemic tissues, including the intestines, blood, thymus, mesenteric lymph nodes, and liver. These experiments identified milk-derived fluorescent signals in intestinal epithelial and immune cells as well as liver macrophages in 7-day-old pups. However, the minute numbers of macrophages in mouse milk suggest that maternal cells are not systemically accumulating in the infant; instead, pup macrophages are consuming milk cell membrane components, such as apoptotic bodies or extracellular vesicles (EVs). Exvivo experiments using primary macrophages support this hypothesis, showing that immune cells preferentially consumed EVs over milk cells. Together, these data suggest a more complex interplay between milk cells and the infant's immune and digestive systems than previously recognized and highlight the need for future research on the role of milk cells in infant health.

The Role of Small Extracellular Vesicles Derived from Glial Cells in the Central Nervous System under both Normal and Pathological Conditions.

In recent decades, researchers and clinicians have increasingly focused on glial cell function. One of the primary mechanisms influencing these functions is through extracellular vesicles (EVs), membrane-bound particles released by cells that are essential for intercellular communication. EVs can be broadly categorized into four main types based on their size, origin, and biogenesis: large EVs, small EVs (sEVs), autophagic EVs, and apoptotic bodies. Small EVs (sEVs) are involved in various physiological and pathological processes such as immune responses, angiogenesis, and cellular communication, primarily by transferring proteins, lipids, and nucleic acids to recipient cells. Interactions among glial cells mediated by small EVs can significantly modulate cell polarization and influence glial behavior through miRNA transfer. This communication, facilitated by small EVs in glial cells, is crucial for neuroinflammation, immune responses, and disease progression. This comprehensive review focuses on driven by glial small EVs, highlighting their roles in transporting biomolecules and modulating the functions of recipient cells. Furthermore, we provide an in-depth overview of the specific contributions of small EVs derived from three principal types of glial cells: oligodendrocytes, astrocytes, and microglia.

The Extra-Tumoral Vaccine Effects of Apoptotic Bodies in the Advancement of Cancer Treatment.

The induction of apoptosis in tumor cells is a common target for the development of anti-tumor therapies; however, these therapies still leave patients at increased risk of disease recurrence. For example, apoptotic tumor cells can promote tumor growth and immune evasion via the secretion of metabolites, apoptotic extracellular vesicles, and induction of pro-tumorigenic macrophages. This paradox of apoptosis induction and the pro-tumorigenic effects of tumor cell apoptosis has begged the question of whether apoptosis is a suitable cancer therapy, and led to further explorations into other immunogenic cell death-based approaches. However, these strategies still face multiple challenges, the most critical of which is the tumor microenvironment. Contrary to the promotion of immune tolerance mediated by apoptotic tumor cells, apoptotic bodies with enriched tumor-related antigens have demonstrated great immunogenic potential, as evidenced by their ability to initiate systemic T-cell immune responses. These characteristics indicate that apoptotic body-based therapies could be ideal "in situ" extra-tumoral tumor vaccine candidates for the treatment of cancers, and further address the current issues with apoptosis-based or immunotherapy treatments. Although not yet tested clinically, apoptotic body-based vaccines have the potential to better treatment strategies and patient outcomes in the future.

Microvesicles Released by Osteoclastic Cells Exhibited Chondrogenic, Osteogenic, and Anti-Inflammatory Activities: An Evaluation of the Feasibility of Their Use for Treatment of Osteoarthritis in a Mouse Model

Extracellular vesicles (EVs), particularly exosomes (EXOs) of various skeletal and stem cells, were shown to delay osteoarthritis (OA) progression, and apoptotic bodies (ABs), another EV subtype, of osteoclasts showed osteoanabolic actions and were involved in the osteoclastic-regulation of local bone formation. Moreover, this study demonstrates that microvesicles (MVs) released by osteoclasts displayed potent pro-chondrogenic, pro-osteogenic, and anti-inflammatory activities. These activities were unique to osteoclastic MVs and were not shared by osteoclastic ABs and EXOs or MVs of other cell types. Because chronic synovial inflammation, progressive articular cartilage erosion, abnormal subchondral bone remodeling, and inability to regenerate articular cartilage are key etiologies of OA, we postulate that the foregoing activities of osteoclastic MVs could simultaneously target multiple etiologies of OA and could thereby be an effective therapy for OA. Accordingly, this study sought to assess the feasibility of an osteoclastic MV-based strategy for OA with a mouse tibial plateau injury model of OA. Briefly, tibial plateau injuries were created on the right knees of adult C57BL/6J mice, MVs were intraarticularly injected into the injured joints biweekly, and the OA progression was monitored histologically at five weeks post-injury. The MV treatment reduced the OA-induced losses of articular cartilage area and thickness, decreased irregularity in the articular cartilage surface, reduced loss of gliding/intermediate zone of articular cartilage, reduced osteophyte formation, suppressed synovial inflammation, and decreased the OARSI OA score. In summary, treatment with osteoclastic MVs delayed or reversed OA progression. Thus, this study supports the feasibility of an osteoclastic MV-based therapy for OA.

Jujuboside B inhibits proliferation and induces apoptosis and ferroptosis in colorectal cancer cells with potential involvement of the MAPK signaling pathway.

Colorectal cancer (CRC) is one of the most prevalent and life-threatening malignancies worldwide. Jujuboside B (JUB) is a bioactive compound derived from the seeds of Ziziphus jujuba, known for its potential anticancer properties. The present study aimed to investigate the association of JUB with inhibiting the proliferation, apoptosis and ferroptosis of human CRC cells with mitogen-activated protein kinase (MAPK) pathway regulation. First, the human CRC HCT116 cell line was treated with different concentrations of JUB. Subsequently, cell viability was evaluated using MTT assay and colony formation was assessed using a colony formation assay. Flow cytometry was used to detect cell apoptosis and the levels of reactive oxygen species. Western blotting was utilized to assess the expression levels of apoptosis-related proteins, ferroptosis regulators and MAPK pathway-related proteins. In addition, biochemical assay kits were used to evaluate the levels of malondialdehyde, glutathione, total iron and ferrous iron. The results demonstrated that cell viability and colony formation were markedly decreased after JUB treatment, whilst the level of apoptosis was notably increased in a concentration-dependent manner. Using electron microscopy, cells treated with JUB exhibited typical apoptotic bodies, as well as mitochondrial swelling and cristae disruption, further demonstrating JUB-induced cell apoptosis. Western blot analysis indicated that JUB treatment markedly reduced the expression of B-cell lymphoma-2 (Bcl-2) but notably increased the expression of Bcl-2 associated X-protein and cleaved caspase-3. Additionally, JUB induced ferroptosis and inhibited the MAPK signaling pathway in CRC cells. Collectively, the findings of the present study suggest that JUB has the potential to inhibit CRC cell proliferation and induce apoptosis through regulating the MAPK pathway. Therefore, JUB may be a promising therapeutic agent for the treatment of CRC.

In situ vaccine "seeds" for enhancing cancer immunotherapy by exploiting apoptosis-associated morphological changes.

Despite the development of many effective immunoadjuvants (IAs), the therapeutic efficacy of in situ vaccines for anti-tumor applications remains limited. Inspired by the morphological changes occurring during apoptosis, this study aims to leverage the release process of autologous tumor antigens (ATAs) to enhance the anti-tumor activity of in situ vaccines. We developed five distinct liposomes, each with unique characteristics and functions, incorporating FDA-approved monophosphoryl lipid A (MPLA) adjuvants into their lipid bilayers. Our findings revealed that the apoptotic bodies generated from tumor cells treated with membrane-fusion liposomes (MFLs) exhibited a greater capacity for immune activation. Mechanistic studies demonstrated that MFLs can utilize the morphological changes associated with apoptosis to accurately deliver adjuvants to apoptotic bodies. To further optimize the efficiency of antigen presentation by these apoptotic bodies as an adjuvant redistribution platform, we encapsulated a calcium chelator within the MFLs to inhibit the externalization of phosphatidylserine (PS) during apoptosis. Through a series of apoptosis-related cellular events, the vaccine can widely disseminate immunoadjuvants (IAs) within tumor tissues, similar to the dispersal of plant seeds. To the best of our knowledge, this is the first approach to utilize apoptosis-associated morphological changes to enhance the immunotherapeutic efficacy of cancer vaccines.

NO-mediated DNA damage induced by polystyrene nanoparticles triggers program cell death in mesenchymal stem cells

Daily contact with considerable amounts of polystyrene nanoparticles (PSNPs) may cause harmful effects on the living organisms, through mechanisms that are not fully understood. The study aimed to evaluate the cytotoxic and genotoxic effects of PSNPs (size 200 nm and 40 nm) in mesenchymal stem cells (MSCs). In order to estimate cellular uptake and retention of nanoplastics, PSNP-treated cells have been analyzed by transmission electron microscopy. For assessing morphology and viability of MSCs after PSNP treatment at two environmentally relevant doses (0.47 and 4.7 μl/ml) for 24 hours, HE and Giemsa staining were performed. Annexin V‑FITC/PI assay was used to quantify PSNPs-mediated cell death. Genotoxicity of PSNPs was evaluated by Comet test. The capacity of PSNPs to trigger the production of free radicals in MSCs was also evaluated. TEM confirmed endocytosis of PSNPs. Decreased cell volume, nuclear hyperchromatism, edge aggregation, and formation of densely stained apoptotic bodies, indicated that PSNP-treated MSCs undergo apoptosis. The presented data showed that both concentration of PS particles significantly increased early apoptotic cell death in comparison to untreated cells. Moreover, both doses of PSNPs significantly increased the genetic damage index in MSCs in dose-dependent manner. In conclusion, PSNPs penetrate, accumulate and induce cytotoxic and genotoxic damage in MSCs.

Preparation of Small Extracellular Vesicles Using Sequential Ultrafiltration with Regenerated Cellulose Membranes of Different Molecular Weight Cutoffs: A Study of Morphology and Size by Electron Microscopy.

There is still room for improvement in the isolation and purification techniques for extracellular vesicles (EVs), particularly in the separation of exosomes (small EVs) from other membrane vesicles such as microvesicles and apoptotic bodies. Furthermore, it is crucial to establish preparation methods that preserve the intrinsic properties of EVs in this context. In this study, we focus on the isolation and preparation of small EVs, exosomes, from the culture supernatant of a human cell line. We discuss the sequential use of regenerated cellulose membranes with different molecular weight cutoffs, based on direct evaluation by transmission electron microscopy, and examine the challenges of characterizing biological membrane vesicles, small EVs, identified during this process.

Biosynthesis of Lysosomally Escaped Apoptotic Bodies Inhibits Inflammasome Synthesis in Macrophages.

Hyperglycemia and bacterial colonization in diabetic wounds aberrantly activate Nod-like receptor protein 3 (NLRP3) in macrophages, resulting in extensive inflammatory infiltration and impaired wound healing. Targeted suppression of the NLRP3 inflammasome shows promise in reducing macrophage inflammatory disruptions. However, challenges such as drug off-target effects and degradation via lysosomal capture remain during treatment. In this study, engineered apoptotic bodies (BHB-dABs) derived from adipose stem cells loaded with β-hydroxybutyric acid (BHB) were synthesized via biosynthesis. These vesicles target M1-type macrophages, which highly express the folic acid receptor in the inflammatory microenvironment, and facilitate lysosomal escape through 1,2-distearoyl-sn-propyltriyl-3-phosphatidylethanolamine-polyethylene glycol functionalization, which may enhance the efficacy of NLRP3 inhibition for managing diabetic wounds. Invitro studies demonstrated the biocompatibility of BHB-dABs, their selective targeting of M1-type macrophages, and their ability to release BHB within the inflammatory microenvironment via folic acid and folic acid receptor signaling. These nanovesicles exhibited lysosomal escape, anti-inflammatory, mitochondrial protection, and endothelial cell vascularization properties. Invivo experiments demonstrated that BHB-dABs enhance the recovery of diabetic wound inflammation and angiogenesis, accelerating wound healing. These functionalized apoptotic bodies efficiently deliver NLRP3 inflammasome inhibitors using a dual strategy of targeting macrophages and promoting lysosomal escape. This approach represents a novel therapeutic strategy for effectively treating chronic diabetic wounds.

Canagliflozin inhibits hedgehog interacting protein (Hhip) induction of tubulopathy in diabetic Akita mice.

Renal hedgehog interacting protein (Hhip) activates sodium-glucose cotransporter 2 (Sglt2) expression and promotes tubular senescence in murine diabetic kidney disease (DKD), yet its underlying mechanism(s) are poorly understood. Here we study the effect of the SGLT2 inhibitor, canagliflozin on tubulopathy (fibrosis and apoptosis) in Akita/HhipRPTC-transgenic (Tg) mice with overexpression of Hhip in their renal proximal tubular cells (RPTCs) and its relevant mechanisms. The DKD-tubulopathy with pronounced Sglt2 expression was aggravated in the kidney of Akita/HhipRPTC-Tg cf. Akita/non-Tg mice. A strong association was observed between Hhip and tubular senescence in Nephroseq from the Nakagawa chronic kidney disease study. Both in vivo and in vitro, excessive Hhip in RPTCs triggered RPTC senescence (polyploidization and cytoskeleton destabilization) and released extracellular vesicles (EVs) carrying Hhip (EVsHhip), most of which were apoptotic bodies (ABsHhip) or microvesicles (MVsHhip) and little exosomes (EXOsHhip). Further, Hhip stimulated β2-microglobulin, which further interacts with EVsHhip, together facilitating RPTC turn-over from cellular senescence to fibrosis and/or apoptosis, ultimately leading to advanced tubulopathy. In contrast, canagliflozin administration offset the action of Hhip in RPTCs, thereby preventing DKD progression. In conclusion, canagliflozin prevented excessive Hhip-mediated tubulopathy, possibly via the inhibition of excessive Hhip carried by extracellular vehicles in DKD.

Zinc Supplementation Mitigates High Salt Diet-Induced Bone Damage: A Histological Evaluation of Osteocyte Apoptosis

Objective: This study aims to evaluate the protective role of zinc supplementation against bone damage induced by a high-salt diet, with a specific focus on osteocyte apoptosis in rats. Methods: This was an investigational study conducted at the Army Medical College, Rawalpindi, Pakistan, over eight weeks. Thirty female Sprague-Dawley rats were randomly divided into three groups: control (Group C), high-salt diet (Group A), and high-salt diet with zinc supplementation (Group B). Group C received a standard diet, Group A received a diet with 8% sodium chloride, and Group B received the high-salt diet plus zinc supplementation at 50 mg/kg body weight daily via oral gavage. After eight weeks, femurs were harvested, processed, and stained with hematoxylin and eosin. Osteocyte apoptosis was assessed by counting empty lacunae and apoptotic bodies under light microscopy. Results: The high-salt diet group (Group A) exhibited a significantly higher density of apoptotic osteocytes compared to the control group (mean ± SD: 2.3166 ± 0.820 vs. 1.3666 ± 0.431 per unit area; p=0.0005). Zinc supplementation in Group B resulted in a significant reduction in osteocyte apoptosis compared to both the high-salt diet group (mean ± SD: 1.7000 ± 0.492 per unit area; p=0.0087) and the control group (p=0.0009). These findings indicate that zinc supplementation effectively reduces osteocyte apoptosis caused by high salt intake. Conclusion: Zinc supplementation significantly mitigates the harmful effects of a high-salt diet on bone health by reducing osteocyte apoptosis. This suggests its potential as a therapeutic intervention to counteract salt-induced bone damage and prevent related diseases such as osteoporosis. Keywords: Zinc, salt, osteocytes, apoptosis, osteoporosis.

Pink1/Parkin signaling mediates pineal mitochondrial autophagy dysfunction and its biological role in a comorbid rat model of depression and insomnia

Using a chronic unpredictable mild stress (CUMS) combined with multi-platform water environment sleep deprivation (SD) as an animal model, the occurrence and development of human depression combined with insomnia were simulated. The abnormal mitochondrial autophagy signaling caused by the putative kinase 1/Parkin E3 ubiquitin protein ligase (Pink1/Parkin) signaling pathway directly affects the normal secretion of melatonin by the pineal gland, which may explain the pathogenesis of depression combined with insomnia. This study aims to explore the depression-like behavior, sleep changes, central oxidative stress response, pineal mitochondrial autophagy damage, melatonin secretion, histopathological changes of the pineal gland, and the expression of Pink1/Parkin signaling-related factors in CUMS+SD rats. The results showed that the levels of reactive oxygen species (ROS) in cerebrospinal fluid of CUMS+SD rats significantly increased along with the inflammatory factors Interleukin-1β (IL-1β) and nuclear factor kappa-B (NF-κB) in cerebrospinal fluid. In addition, the number of pineal gland cells significantly decreased, cell boundaries became blurred, cell volume shrank, and apoptotic bodies appeared in the pineal gland tissue under HE staining, indicating pineal gland inflammation. Sleep deprivation further disrupted the levels of autophagy damage factors, including histamine (MDA), glutathione (GSH), and catalase (CAT), in the cerebrospinal fluid of CUMS + SD rats. Transmission electron microscopy of the pineal gland in CUMS+SD rats revealed damage to mitochondrial autophagy. The levels of 5-hydroxytryptamine (5-HT) and aromatic amine-N-acetyltransferase (AANAT) in the cerebrospinal fluid, as well as melatonin levels in the pineal gland, were significantly decreased. Additionally, the expression of IL-1β, NF-κB, Pink1, and Parkin in the pineal gland of CUMS+SD rats significantly increased. The expression of microtubule-associated protein 1 light chain 3-β (LC3), selective autophagy adaptor protein (P62), cytochrome c oxidase IV (COXIV), and mitochondrial outer membrane translocation enzyme 20 (TOM20) proteins downstream of the Pink1/Parkin signaling pathway was enhanced, while the expression of downstream brain-derived neurotrophic factor (BDNF), Beclin 1, and BCL2 interacting protein 3 (BNIP3) proteins was negatively regulated. Pink1/Parkin signaling may specifically respond to mitochondrial autophagy damage in the pineal gland, affecting the normal synthesis and secretion of melatonin in the pineal gland. In summary, mitochondrial autophagy damage in the pineal gland affects the normal secretion of melatonin in CUMS+SD rats, which is closely related to the specific autophagy signaling impairment of Pink1/Parkin pathway, which may mediate the occurrence of depression combined with insomnia.

Extracellular vesicles: from intracellular trafficking molecules to fully fortified delivery vehicles for cancer therapeutics.

Extracellular vesicles (EVs) are emerging as viable tools in cancer treatment due to their ability to carry a wide range of theranostic activities. This review summarizes different forms of EVs such as exosomes, microvesicles, apoptotic bodies, and oncosomes. It also sheds the light onto isolation methodologies, characterization techniques and therapeutic applications of all discussed EVs. Evidence indicates that EVs are particularly effective in delivering chemotherapeutic medications, and immunomodulatory agents. However, the advancement of EV-based therapies into clinical practice is hindered by challenges including EVs heterogeneity, cargo loading efficiency, and in vivo stability. Overall, EVs have the potential to change cancer therapeutic paradigms. Continued research and development activities are critical for improving EV-based medications and increasing their therapeutic impact.

Visible-light-induced CO-releasing properties and cytotoxicity of a Ru(II) carbonyl complex containing 2-(pyridin-2-yl)-quinoxaline.

The photo-induced CO-releasing properties of the dark-stable complex [RuCl2(CO)2L] (L = 2-(pyridin-2-yl)quinoxaline) were investigated under 468 nm light exposure in the presence and absence of biomolecules such as histidine, calf thymus DNA and hen egg white lysozyme. The CO release kinetics were consistent regardless of the presence of these biomolecules, suggesting that they did not influence the CO release mechanism. The quinoxaline ligand demonstrated exceptional cytotoxicity against human acute monocytic leukemia cells (THP-1), with evidence of potential DNA damage ascertained by comet assay, while it remained non-toxic to normal kidney epithelial cells derived from African green monkey (Vero) cell lines. In contrast, upon light activation, the Ru(II) complex showed no toxicity against THP-1 cells but was detrimental to Vero cells. In human colorectal carcinoma (HCT-116) cells, the ligand and the Ru(II) complex produced ROS under light and dark conditions. However, HCT-116 cells retained their ability to consume oxygen and produce ATP following CO treatment, suggesting that the ROS levels were insufficient to cause significant cellular damage. Morphological features of apoptosis, including apoptotic bodies, chromatin condensation, cell shrinkage, and membrane leakage, were observed in the presence of both the ligand and its complex, irrespective of light exposure.

External pressure induced the dysfunction of Sertoli cells via the Fas/FasL signaling pathway

Cryptorchidism, a condition where the testis fails to fully descend into the scrotum during development, is associated with elevated environmental temperatures and pressures, leading to male infertility and germ cell tumors. Factors such as oxidative stress and high temperatures contribute to infertility in cryptorchidism. This study aims to explore how external pressure affects Sertoli cells and discover new mechanisms affecting spermatogenesis in cryptorchidism. Sertoli cells were subjected to various pressure levels (0 mmHg, 25 mmHg, 50 mmHg, 100 mmHg) and durations (0 h, 2 h, 4 h) using an enzyme-linked immunosorbent assay (ELISA) to measure androgen binding protein (ABP) and inhibin B (INH B) secretion. Cell morphology changes were observed using immunofluorescence; apoptosis rates were measured with terminal-deoxynucleotidyl transferase mediated nick end labelling (TUNEL) assay and flow cytometry; ultrastructural variations were examined via transmission electron microscopy; and the expression of apoptosis-related proteins (Fas, FasL, caspase 3, and caspase 8) was analyzed through immunohistochemistry, real-time polymerase chain reaction (real-time PCR), and western blotting. The results showed that elevated pressure suppressed ABP and INH B secretion from Sertoli cells. Structural changes were observed under pressure, including cytoskeleton loosening and nuclear fragmentation. Apoptosis rates increased with higher pressure levels. Ultrastructural analysis revealed chromatin changes, apoptotic bodies, and mitochondrial alterations. Increased expressions of Fas and FasL were detected, along with elevated levels of caspase 3 and caspase 8. The caspase 8 inhibitor blocked pressure-induced apoptosis and caspase 3 activation, while the cytochrome C inhibitor did not show the same effect. Our findings suggested that external pressure induces apoptosis of Sertoli cells via the Fas/FasL signaling pathway, potentially contributing to male infertility associated with cryptorchidism.

Apoptotic body based biomimetic hybrid nanovesicles to attenuate cytokine storms for sepsis treatment

Sepsis is a severe immune response to pathogens that is associated with high mortality rate and a paucity of efficacious treatment options. It is characterized by the hyperactivation of macrophages and the occurrence of cytokine storms. Given the anti-inflammatory properties of M2 macrophages and their derived apoptotic bodies (AB), as well as the specific uptake of these by macrophages, a novel approach was employed to combine AB with artificial liposomes to create apoptotic body based biomimetic hybrid nanovesicles (L-AB). The L-AB effectively inherited “eat me” signaling molecules on the surface of the AB, thereby facilitating their targeted uptake by macrophages in both in vitro and in vivo settings. The administration of L-AB for the delivery of dexamethasone effectively augmented the therapeutic efficacy of the drug, mitigated macrophage hyperactivation and tissue damage in vivo, and consequently enhanced the survival rate of septic mice. Taken together, these findings suggest that the apoptotic body biomimetic nanovesicles may represent a potential drug delivery system capable of specifically targeting macrophages for the treatment of sepsis.

Neutrophil-derived apoptotic body membranes-fused exosomes targeting treatment for myocardial infarction.

Myocardial infarction (MI) poses a substantial threat to human health, prompting extensive research into effective treatment modalities. Preclinical studies have demonstrated the therapeutic potential of mesenchymal stem cell-derived exosomes for cardiac repair. Despite their promise, the inherent limitations of natural exosomes, mainly their restricted targeting capabilities, present formidable barriers to clinical transformation. To address this, it is proposed to enhance their targeting specificity and retention in infarcted myocardium by fusing exosomes with neutrophil-derived apoptotic body membranes (NAM). These NAM inherit the surface signals from neutrophils, which allow them to home in on the damaged tissues and participate in regulating inflammatory responses. In this current work, we utilized a membrane fusion technique to create NAM-fused exosomes (NAM-Exo) for MI treatment. Compared to their native counterparts, NAM-Exo demonstrated enhanced adhesion to inflammatory endothelial cells, replicating the neutrophil recruitment mechanism at sites of myocardial injury in MI. Furthermore, our findings revealed that NAM-Exo not only significantly modulated inflammation responses but also promoted angiogenesis in a mouse model of MI, ultimately leading to improved cardiac function and ventricular remodeling post-treatment. These results underscore the potential of membrane fusion as an effective strategy to enhance the therapeutic efficacy of exosome-based cardiac repair and regeneration therapies, thereby paving the way for their translation into clinical practice.

Integrating eugenol with intensive care in leukemia patients: exploration of pro-apoptotic potential against HL-60, human leukemia cell line

Objective: The objective of this in-vitro study was to explore the pro-apoptotic potential of eugenol (4-allyl-2-methoxyphenol) on Human Leukemia-60 (HL-60) cell line as a potent phytochemical in intensive care setting to leukemia patients. Methodology: After formal approval by all of the respective ethical committees, this study was simultaneously conducted at all respective institutional departments. The study included culturing HL-60 cell line and its treatment with serial concentrations of eugenol for calculation of subsequent IC50 values (half-maximal inhibitory concentration) via MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay, expression analysis of gene, comparative analysis of relative gene fold (Intrinsic Biomarkers Caspase-3, Caspase-9) as apoptosis mediator markers followed by RT-qPCR. Cellular apoptotic morphology was confirmed via Hoechst 333258 staining. Results: For HL-60 cell line, the IC50 of eugenol (14.1 uM) showed high gene expression of pro-apoptotic biomarkers (Caspase-3 and Caspase-9). Hoechst 333258 staining showed prominent apoptotic bodies leading to nuclear fragmentations. Conclusion: Eugenol proved to possess robust pro-apoptotic potential leading to diagnostic efficacy against leukemia HL-60 cell line. Further studies would help in identifying key mechanisms by which eugenol exhibits anti-cancer potential against HL-60 cell lines. Abbreviations: HL-60 - Human Leukemia-60; IC50 - Half-Maximal Inhibitory Concentration Keywords: Eugenol, Pro-apoptotic Effects, Intensive Care, Anti-Cancer Mechanisms, HL-60, Natural Chemotherapeutics Citation: Khaliq HMH, Bughio R, Nangdev P, Aziz O, Javed W. Integrating eugenol with intensive care in leukemia patients: exploration of pro-apoptotic potential against HL-60, human leukemia cell line. Anaesth. pain intensive care 2024;28(5):871−875; DOI: 10.35975/apic.v28i5.2491 Received: June 18, 2024; Reviewed: August 03, 2024; Accepted: August 11, 2024