Necrotic Features Research Articles

PANoptosis in autoimmune diseases interplay between apoptosis, necrosis, and pyroptosis

PANoptosis is a newly identified inflammatory programmed cell death (PCD) that involves the interplay of apoptosis, necrosis, and pyroptosis. However, its overall biological effects cannot be attributed to any one type of PCD alone. PANoptosis is regulated by a signaling cascade triggered by the recognition of pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) by various sensors. This triggers the assembly of the PANoptosome, which integrates key components from other PCD pathways via adapters and ultimately activates downstream execution molecules, resulting in cell death with necrotic, apoptotic, and pyroptotic features. Autoimmune diseases are characterized by reduced immune tolerance to self-antigens, leading to abnormal immune responses, often accompanied by systemic chronic inflammation. Consequently, PANoptosis, as a unique innate immune-inflammatory PCD pathway, has significant pathophysiological relevance to inflammation and autoimmunity. However, most previous research on PANoptosis has focused on tumors and infectious diseases, leaving its activation and role in autoimmune diseases unclear. This review briefly outlines the characteristics of PANoptosis and summarizes several newly identified PANoptosome complexes, their activation mechanisms, and key components. We also explored the dual role of PANoptosis in diseases and potential therapeutic approaches targeting PANoptosis. Additionally, we review the existing evidence for PANoptosis in several autoimmune diseases and explore the potential regulatory mechanisms involved.

The utility of PET-CT in baseline and sequential characterisation of Pulmonary Pleomorphic Carcinoma

Pulmonary Pleomorphic Carcinomas (PPCs) represent a rare and aggressive subtype of Non-Small Cell Lung Cancer (NSCLC) that can only be definitively diagnosed on a surgical specimen. This study utilised PET-CT to evaluate radiological characteristics of PPCs. This study retrospectively evaluated the radiological characteristics of PCCs diagnosed in St James’s Hospital Dublin between 2012-2023. Computed Tomography (CT) and Positron Emission Tomography (FDG-PET) imaging features (size, location, density, shape, invasion, and growth kinetics) and standard uptake value for each lesion were evaluated. 39 PCCs were identified with a mean age of 66.5 years (range: 49-82 years). FDG-PET was performed in all 39 cases. Tumours demonstrated a high FDG uptake at baseline with a mean (SUV) of 12.6 (range: 1.4 - 36.9). A second interval PET-CT on average 3.3 months after the first in 3 cases demonstrated over 120% increase in SUV. The mean tumour size was 4.3 cm (range: 1.0 - 14.5 cm). Tumours developed rapid interval growth, reaching a mean maximum diameter of 6.6 cm (53.4%) within a mean of 2.1 months. Tumours were predominantly located in the upper lobe (71.8 %) and displayed necrotic features in 53.8 % of cases. 82.1% of tumours invaded the mediastinum. This study describes the largest cohort of Pulmonary Pleomorphic Carcinoma in the literature. Tumours demonstrate a high SUV on baseline imaging and demonstrate rapid growth on interval imaging and central necrosis. Please click on the 'PDF' for the full abstract!

The in vitro cytotoxic effects of natural (fibrous epsomite crystals) and synthetic (Epsom salt) magnesium sulfate.

Exposure to mineral fibers represents an occupational and environmental hazard since particulate inhalation leads to several health disorders. However, few data are available on the effect of fibers with high solubility like natural epsomite, a water-soluble fiber with an inhalable size that allows it to penetrate biological systems, with regard to the respiratory tract. This study evaluated the natural (fibrous epsomite) and synthetic (Epsom salt) magnesium sulfate pathogenicity. Investigations have been performed through morpho-functional and biochemical analyses, in an in vitro cell model that usually grows as monocytes, but that under appropriate conditions differentiates into macrophages. These latter, known as alveolar macrophages, if referred to lungs, represent the first line of defense against harmful inhaled stimuli. Morphological observations reveal that, if Epsom salt induces osmotic stress on cell culture, natural epsomite fibers lead to cellular alterations including thickening of the nuclear envelope and degenerated mitochondria. Moreover, the insoluble fraction (impurities) internalized by cells induces diffuse damage characterized at the highest dosage and exposure time by secondary necrosis or necrotic cell death features. Biochemical analyses confirm this mineral behavior that involves MAPK pathway activation, resulting in many different cellular responses ranging from proliferation control to cell death. Epsom salt leads to MAPK/ERK activation, a marker predictive of overall survival. Unlike, natural epsomite induces upregulation of MAPK/p38 protein involved in the phosphorylation of downstream targets driving necrotic cell death. These findings demonstrate natural epsomite toxicity on U937 cell culture, making the inhalation of these fibers potentially hazardous for human health. RESEARCH HIGHLIGHTS: Natural epsomite and synthetic Epsom salt effects have been evaluated in U937 cell model. Epsom salt induces an osmotic cellular stress. Natural epsomite fibers lead to cellular damage and can be considered potentially dangerous for human health.

Oncolytic NDV-AMHA1 Strain Effective Against Glioblastoma Cancer Cells

Glioblastoma multiforme (GBM) is one of the most life-threatening types of cancer that is difficult to treat. The search for effective yet safe therapy is progressing and non-conventional therapies such as using viruses as a smart and selective agent against cancer are promising. The aim of the study was the presence of a reliable method to use Newcastle disease virus (NDV) as an oncolytic agent against GBM, which attempted to propagate the NDV in laboratory experiments. Ahmed Majeed Hamza Al-Shammari-1 (AMHA1) attenuated strain of NDV was propagated in chicken embryos. The virus's tittered in Vero-slamed cells to determine the infective dose. MTT cell viability assay was used to investigate the killing effects of NDV on Ahmed-Majeed-Glioblastoma-Multiforme-2005 (AMGM5) human glioblastoma cancer cells derived from Iraqi patients. The infected cells' morphology was studied to measure the cytolytic effect of the NDV in cancer cells. Results showed that After 24 to 72 hours of inoculation, all of the chicken embryos died when the AMHA1 Iraqi NDV strain was injected. Cell viability assay showed that the NDV-AMHA1 strain has cytotoxicity at MOI of 0.1, 0.5, and 1 for 72 hours of exposure to cancer cells. The morphological analysis showed that NDV induces cell death in the infected cells with both necrotic and apoptotic features. In conclusion, the study focuses on the propagation of the oncolytic NDV as a biological agent capable of overcoming treatment resistance through infecting and replicating inside cancer.

RETRACTED: The heterogeneity of PD-L1 protein in gastric cancer: expression and distribution characteristics

Programmed death receptor ligand 1 (PD-L1) is expressed at different levels in tumour tissues and tumour-infiltrating monocytes (TIMCs). The interpretation of PD-L1 expression in gastric cancer (GC) is more difficult because of its heterogeneity. The PD-L1 immunohistochemistry (IHC) by E1L3N assay was performed in GC tissues. The level and distributed characteristics of PD-L1 expression were observed to illustrate its heterogeneity both in the cancer tissues and TIMCs. The relationship between PD-L1 level and necrotic features of tumor cells, the number of TIMCs, the distribution of tertiary lymphoid tissue (TLS) in the stroma, and other clinicopathological factors were analysed. A Cox regression model was used to assess the prognostic value of PD-L1 expression. Of the 110 GC samples, not only more cases (51/110 cases) could be detected by combined positive score (CPS) for PD-L1 expression compared the other two, tumour positive score (TPS), and mononuclear immune-cell density score (MIDS), but also there were more cases with the high level of PD-L1 expression by CPS, even if with good consistency among them (P < 0.05). The tumour cells with high expression of PD-L1 was prone to show a diffuse distributing, whereas mottled type in the low level. It was noteworthy that the strongly colored tumor cells tended to exhibit a mossy pattern which were distributed along the border between cancer nests and stroma, and the same pattern happened to occur in the positive mesenchymal cells contacting the tumor border, essentially lymphocytes and macrophages. The substantial necrosis in the tumour and the number of TIMCs was analyzed statistically significant correlated with CPS (P < 0.05), while other clinicopathological factors such as histological type, tumour size, invasion depth, TNM stage were uncorrelated. The number and distribution of TLS in the tumour and para-tumoural stroma indirectly affected PD-L1 in GC by associating with the quantity and pattern of TIMCs. Cox regression analysis revealed that the prognosis was poor when PD-L1 was positive. CPS is the best indicator for PD-L1 expression in GC, which tend to be increased expression following a large number of TIMCs and substantial tumour necrosis appeared. Heterogeneity was reflected in the different distributed pattern of PD-L1 expression, especially the mossy-like pantten of the staining tumor cell in the interface between tumour nests and stroma, regardless of the amount and intensity of PD-L1 expression. TLS is valuable for observing microscopic images to influence the quantity and pattern of TIMCs. CPS can be used as an independent prognostic factor for GC.

Characterization of green route synthesized zinc oxide nanoparticles using Cyperus rotundus rhizome extract: Antioxidant, antibacterial, anticancer and photocatalytic potential

Cyperus rotundus rhizome has been employed in traditional Chinese and Indian Ayurvedic herbal formulations. Green synthesis of metal oxide nanoparticles has gained significant attention due to its economic, biocompatibility, non-toxicity and eco-friendliness. The current study confirmed the biosynthesized zinc oxide nanoparticles (ZnONPs) by an ultraviolet–visible absorption spectrum of 353 nm. Fourier transform infrared spectroscopy analysis demonstrated that bioactive functional groups served as reducing and stabilizing agents for ZnONPs. Further, X-ray powder diffraction (XRD) patterns showed crystalline nature with a hexagonal phase and strongly correlated with JCPDS file number 36-1451. The electron microscopic studies revealed irregular spherical particles with a size range of 17–100 nm and EDAX analysis validated the crystalline purity of ZnONPs. The average size of the ZnONPs determined by dynamic light scattering was 95.89 nm and the zeta potential was −12.1 mV, indicating stability of the particles. Moreover, the ZnONPs exhibited noticeable antioxidant potential by scavenging free radicals on DPPH (59.74%), ABTs (66.35%), nitric oxide (65.75%) and reducing power (63.94%) in a dose-dependent manner. The bacterial cell damages were observed through FESEM, confirming the antibacterial efficacy of ZnONPs. Further, the ZnONPs exhibited remarkable cytotoxic effect (62.34 μg/mL) against breast cancer cells and subsequent fluorescence staining demonstrated apoptotic and necrotic features in damaged cells. The photocatalytic effectiveness of ZnONPs demonstrated increased eco-bioremediation capabilities by degrading crystal violet (80.69%) and methylene blue (88.93%) dyes. Thus, this is the first report on ZnONPs synthesized using C. rotundus could be explored as a promising material for biological and photocatalytic applications.

34535 Pseudoherpetic transient acantholytic dermatosis

A 70-year-old woman with newly diagnosed AML awaiting induction chemotherapy presented with pruritic erythematous papulovesicles on her back, abdomen, and extremities. Vesicle fluid HSV and VZV PCR tests were negative, but the patient was diagnosed with presumed disseminated zoster and started on IV acyclovir. However, after 8 days of treatment the rash was not improved. Biopsy at that time revealed hyperkeratosis, focal intraepidermal split with necrotic epidermis, acantholysis, and features consistent with herpetic cytopathic changes including multinucleated keratinocytes and nuclear molding. Immunohistochemical stains for HSV and VZV were negative, and repeat lesional fluid PCR for HSV and VZV were also negative. The patient was diagnosed with pseudoherpetic transient acantholytic dermatosis (TAD) or Grover disease. The rash improved with use of triamcinolone 0.1% cream and ammonium lactate lotion. TAD classically presents with pruritic erythematous papules across the trunk and is thought to be brought on by UV radiation, sweat, heat, or persistent fevers. Histology shows focal acantholysis, dyskeratosis, and intraepidermal clefting. Variants that resemble other acantholytic diseases such as Darier disease, pemphigus vulgaris and Hailey-Hailey disease have been described. A rare variant, pseudoherpetic TAD (P-TAD), has been reported to mimic HSV/VZV infection with multinucleated and hypereosinophilic keratinocytes but lacks true cytopathic effect despite multinucleation; HSV/VZV immunohistochemistry is negative. Clinically, PHD features ungrouped vesicles distributed regionally rather than the grouped vesicles classically seen in herpetic infection. Clinicians should consider a diagnosis of P-TAD in patients with suspected herpetic infection with negative confirmatory PCR testing and lack of response to antivirals.

Alpha-lipoic acid-role in improving both reserpine toxicity and paroxetine treatment in the cerebral cortex of albino rats; histological, ultrastructural, immunohistohemical and biochemical studies

BackgroundReserpine is a monoamine depletory drug cause oxidative damage and used to induce depression-like features in rodent model. Paroxetine is an antidepressant drug that exerts its effects by inhibiting dopaminergic neurons although it may exert much pathological damage. Alpha-lipoic acid (ALA) is an endogenous antioxidant co-factor of important enzymatic complexes. The present study was aimed to elucidate the possible protective effect of ALA in the improvement of the deleterious cerebral cortex injury after reserpine and paroxetine treatment. Forty adult male albino rats were equally divided into 5 groups. Group I served as control group orally treated with saline solution all the experiment period. Group II animals orally treated with ALA (200 mg/kg/day) for six weeks. The induction of depression-like features occurred when the rest of animals were intraperitoneally treated with 25 mg/kg of reserpine once daily for consecutive 14 day. Then these animals were divided into; Group III (reserpine group) animals in this group were sacrificed on 15th day. Group IV; reserpine-treated animals were treated with paroxetine (20 mg/kg) daily for 6 weeks. Group V, animals in this group were received paroxetine and ALA daily for 6 weeks.ResultsReserpine-treated rats showed disorganized layers of cerebral cortex with degenerative, apoptotic and necrotic changes. Ultrastructure changes include both pyramidal and granule cells with severe degenerative, necrotic and apoptotic features. The nuclei appeared pyknotic; irregular with chromatin condensation as well as the cytoplasm of these cells contained many degenerated organelles. In addition, a significant increase in total oxidative stress and decrease in total antioxidant capacity, norepinephrine, dopamine and serotonin levels were recorded. The same treatment showed significant decrease in proliferating cell nuclear antigen (PCNA) expression and significant increase in caspase-3 expression in the granule and pyramidal cells. After paroxetine-treatment these parameters were more or less similar to those observed in reserpine-treated ones. While an obvious improvement was appeared when animal treated with both paroxetine and ALA and; all parameters restored its normal features.ConclusionsThis study concluded that; ALA treatment attenuated the cerebral injury induced by reserpine and improved the effects of paroxetine in rats due to its anti-inflammatory, anti-apoptotic and antioxidant activities.

Comparative cytotoxic activity of Convolvulus pluricaulis against human hepatoma cell line (HepG2) and normal cell line (L929) via apoptosis pathways by flow cytometry analysis

BackgroundThe purpose of this study was to use the MTT test to assess the cytotoxic effects of different extracts of Convolvulus pluricaulis leaves in vitro. Convolvulus pluricaulis leaf ethanolic extract has been demonstrated to trigger apoptosis on HepG2 cancer cell lines, implying anti-cancer activity.MethodsThe cells were grown in culture DMEM and incubated with different concentrations of the plant extracts. Survival rates were quantified by MTT assays by 24 h of exposure to (640–20 µg/ml) the PEE, CHCl3E, EAE, ETHE and AQE of the plant, while monitoring changes on cellular shapes by inverted phase contrast microscopy (PEE—petroleum ether extract, CHCl3—chloroform extract, EAE—ethyl acetate extract, ETHE—ethanol extract, AQE—aqueous extract).ResultsThe Convolvulus pluricaulis leaves extract showed IC50 value < 1000 μg/ml on HepG2 and IC50 value > 1000 µg/ml on L-929 cell lines. Hence, Convolvulus pluricaulis leaves extracts are non-toxic against the normal cell line L-929. Treatments with standard as a control exhibited necrotic features in both cell lines. On the basis of these findings, and because a highly effective extract ETHE has a partial polarity, this plant cytotoxicity and apoptotic activities were also investigated. On HepG2 cell line, the ETHE showed higher cytotoxicity activities compared to AQE, EAE, CHCl3E, and PEE extracts (P < 0.0001) with inhibitory concentration IC50 values of 35.873 μg/ml that is < 1000 μg/ml. The percentage of apoptotic cells of ETHE was determined using propidium iodide (PI) staining of DNA fragments by flow cytometry.ConclusionsThe extract of Convolvulus pluricaulis has cytotoxic and apoptotic action against the HepG2 cell line, indicating that it should be studied further for cancer therapies.

Oxidative Stress- and Autophagy-Inducing Effects of PSI-LHCI from Botryococcus braunii in Breast Cancer Cells.

Botryococcus braunii (B. braunii) is a green microalga primarily found in freshwater, reservoirs, and ponds. Photosynthetic pigments from algae have shown many bioactive molecules with therapeutic potential. Herein, we report the purification, characterization, and anticancer properties of photosystem I light-harvesting complex I (PSI-LHCI) from the green microalga B. braunii UTEX2441. The pigment–protein complex was purified by sucrose density gradient and characterized by its distinctive peaks using absorption, low-temperature (77 K) fluorescence, and circular dichroism (CD) spectroscopic analyses. Protein complexes were resolved by blue native-PAGE and two-dimensional SDS-PAGE. Triple-negative breast cancer MDA-MB-231 cells were incubated with PSI-LHCI for all of our experiments. Cell viability was assessed, revealing a significant reduction in a time- and concentration-dependent manner. We confirmed the internalization of PSI-LHCI within the cytoplasm and nucleus after 12 h of incubation. Cell death mechanism by oxidative stress was confirmed by the production of reactive oxygen species (ROS) and specifically superoxide. Furthermore, we monitored autophagic flux, apoptotic and necrotic features after treatment with PSI-LHCI. Treated MDA-MB-231 cells showed positive autophagy signals in the cytoplasm and nucleus, and necrotic morphology by the permeabilization of the cell membrane. Our findings demonstrated for the first time the cytotoxic properties of B. braunii PSI-LHCI by the induction of ROS and autophagy in breast cancer cells.

SERPINB3 (SCCA1) inhibits cathepsin L and lysoptosis, protecting cervical cancer cells from chemoradiation

The endogenous lysosomal cysteine protease inhibitor SERPINB3 (squamous cell carcinoma antigen 1, SCCA1) is elevated in patients with cervical cancer and other malignancies. High serum SERPINB3 is prognostic for recurrence and death following chemoradiation therapy. Cervical cancer cells genetically lacking SERPINB3 are more sensitive to ionizing radiation (IR), suggesting this protease inhibitor plays a role in therapeutic response. Here we demonstrate that SERPINB3-deficient cells have enhanced sensitivity to IR-induced cell death. Knock out of SERPINB3 sensitizes cells to a greater extent than cisplatin, the current standard of care. IR in SERPINB3 deficient cervical carcinoma cells induces predominantly necrotic cell death, with biochemical and cellular features of lysoptosis. Rescue with wild-type SERPINB3 or a reactive site loop mutant indicates that protease inhibitory activity is required to protect cervical tumor cells from radiation-induced death. Transcriptomics analysis of primary cervix tumor samples and genetic knock out demonstrates a role for the lysosomal protease cathepsin L in radiation-induced cell death in SERPINB3 knock-out cells. These data support targeting of SERPINB3 and lysoptosis to treat radioresistant cervical cancers.

CS-6 A case of poorly differentiated chordoma with systemic metastasis

A case report: The patient was a 32-year-old man with diplopia. He was diagnosed as sphenoid sinusitis on MRI by a local doctor and visited an otolaryngologist. MRI showed extensive extension of neoplastic lesions from the clivus to the sphenoid sinus to the anterior ethmoid sinuses, bilateral cavernous sinuses, and the right medial and lateral pterygoid muscles. The right Lebiere’s lymph node was enlarged and thought to be a metastatic site. Based on the rapid growth and extension of the tumor, the patient was referred to the Department of Otolaryngology at our hospital on suspicion of sinonasal carcinoma. The possibility of chordoma could not be denied, so the patient was referred to our department. The patient underwent a joint endoscopic extended transsphenoidal tumor resection. The pathological diagnosis showed mitotic and necrotic features, and the majority of the cells showed highly atypical components without mucous substrate. However, brachyury, a marker for chordoma, was diffusely positive, and there was loss of INI1 (SMARCB1) expression. The final diagnosis was poorly differentiated chordoma. Postoperatively, the tumor in the right cavernous sinus grew rapidly, and the right eye became blind due to obstruction of the superior ophthalmic vein. The patient was treated with Gamma Knife as soon as possible in the hope of local control by high-dose irradiation, and after a total of three irradiations, the residual tumor shrank markedly and symptoms improved, but systemic metastasis occurred in a short period of time and the patient died. The number of cases of poorly differentiated chordoma has been reported rarely (more than 50), and it is more common in children and even rarer in adults. We report this case with a review of the literature.

High-Glucose and Free Fatty Acid-Induced Adipocytes Generate Increasing of HMGB1 and Reduced GLUT4 Expression

Background&#x0D; High-mobility group box 1 protein (HMGB1) is released from necrotic adipocytes into the extracellular milieu as an inflammatory alarmin in obesity. Although the impact of excess nutrient on adipocytes is well known, it is not clear how specific its component drive cell-size and damaged of adipocytes, and how this relates to the risk of insulin resistance.&#x0D; Objectives&#x0D; The aim of this study was to determine HMGB1 level in adipocytes cultures after high glucose and/or FFA exposures and to assess GLUT4 expression. We determined cellular features of adipocytes that correlates to HMGB1 released and insulin resistance.&#x0D; Methods&#x0D; Differentiated adipocytes were exposed to high glucose and/or FFAs for 7 days. ELISA was performed on supernatant to assess the HMGB1 level. Total GLUT4 expression were quantified by immunofluorescense.&#x0D; Results&#x0D; High glucose and FFA-exposed cells have significant increase of HMGB1 level with decreased of cell size and necrotic adipocytes features. The total GLUT4 were reduced in HG-cells (p &lt;0,045), but not in FFA cells. Hypertrophic adipocytes (p &lt;0.05) and slight decrease of GLUT4 expression were showed on HG+FFA exposures with no increase of HMGB1 level. There was a significant correlation between cell size and HMGB1 level (R -0,637, p &lt; 0.026)&#x0D; Conclusion&#x0D; The expression level studies between high glucose, FFA, and a combination of both on adipocytes results strongly suggest that high glucose is more damaging to adipocyte compared to FFA. Nevertheless, the combination of the two causes adipocyte dysfunction with general features of adipose tissue in obesity, suggested it can be used as a hypertrophic adipocytes model to study obesity in vitro.

Infectious complications of EBUS-TBNA: A nested case-control study using 10-year registry data

ObjectivesEndobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) has become a standard procedure, but little is known about its infectious complications. The aim of this study is to evaluate the incidence and risk factors of infectious complications of EBUS-TBNA and its clinical course, including effects on anti-cancer treatment. MethodsThis is a nested case-control study of patients who received EBUS-TBNA and were followed for at least 2 months at Samsung Medical Center from August 2009 to April 2019. Patients with clinical symptoms of infection and correlating chest images were defined as the infection group (n = 33). The controls were randomly selected from patients without infectious complication. Multivariate logistic regression with backward selection was used to identify the risk factors of infectious complications. ResultsOf the 6826 patients, 33 (0.48%) infectious complications were identified, comprising pneumonia (n = 20) and mediastinal infections (n = 13). Target lesions with necrotic features on chest computed tomography (CT) scan (adjusted odds ratio [aOR], 3.08; 95% confidence interval [CI], 1.49–6.40; P = 0.002) and procedures that were performed via the esophagus (aOR, 3.19; 95% CI, 1.47–6.88; P = 0.003) were independently associated with infectious complications. Among patients ultimately diagnosed with cancer, the infection group tended to refuse anti-cancer treatment compared to controls (32/459, 7.0% vs. 5/30, 16.7%; P = 0.066). However, among the patients who received anti-cancer treatment, there was no delay in onset of treatment. ConclusionsInfectious complications of EBUS-TBNA are rare; however, attention should be paid if the target lesion appears necrotic on chest CT or if the procedure is performed via the esophagus. Although it was not conclusive due to its rarity, patients with infectious complications tended not to receive anti-cancer treatment.

Inhibitors of Lipoxygenase and Cyclooxygenase-2 Attenuate Trimethyltin-Induced Neurotoxicity through Regulating Oxidative Stress and Pro-Inflammatory Cytokines in Human Neuroblastoma SH-SY5Y Cells.

Trimethyltin (TMT) is an environmental neurotoxin that mediates dopaminergic neuronal injury in the brain. In this study, we characterized the toxic mechanism and possible protective compounds against TMT-induced neurotoxicity in human dopaminergic neuroblastoma SH-SY5Y cells. Antioxidants such as melatonin, N-acetylcysteine (NAC), α-tocopherol, and allopurinol alleviated TMT toxicity. Apoptosis induced by TMT was identified by altered expression of cleaved caspase-3, Bax, Bcl-2, and Bcl-xL through Western blot analysis. The iron chelator deferoxamine ameliorated the alteration of apoptosis-related proteins through TMT exposure. TMT also induced delayed ultrastructural necrotic features such as mitochondrial swelling and cytoplasmic membrane rupture; NAC reduced these necrotic injuries. Esculetin, meloxicam, celecoxib, and phenidone decreased TMT toxicity. Elevation of the pro-inflammatory cytokines IL-1β, TNF-α, and NF-ĸB and reduction of the antioxidant enzymes catalase and glutathione peroxidase-1 (GPx-1) were induced by TMT and ameliorated by inhibitors of LOX and COX-2 enzymes. Both NMDA and non-NMDA antagonists attenuated TMT toxicity. The free calcium ion modulators nimodipine and BAPTA/AM contributed to neuronal survival against TMT toxicity. Inhibitors of the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin pathway, an autophagy regulator, decreased TMT toxicity. These results imply that TMT neurotoxicity is the chief participant in LOX- and COX-2-mediated apoptosis, partly via necrosis and autophagy in SH-SY5Y cells.

Ultrastructural Alterations of Platelets and Monocytes Associated with Autoantibodies in Systemic Lupus Erythematosus

Introduction. Systemic lupus erythematosus (SLE) is a chronic autoimmune disease with a high titer of autoantibodies, and SLE patients are predisposed to thrombosis. Monocytes and platelets are potentially involved in the pathogenesis of SLE. Monocytes are a source of immune modulators for T and B lymphocytes and they increase the risk of thrombosis through exposing tissue factor on their cell surface. Platelets are activated in SLE and may promote thrombotic events by expressing the procoagulant phosphatidylserine. The aim of this work was to investigate the ultrastructural changes of monocytes and platelets in SLE patients in association with titers of autoantibodies.Methods. SLE patients were included in the study based on the diagnostic criteria of the American College of Rheumatology. Platelets from 5 SLE patients and 3 healthy donors were isolated by gel-filtration on Sepharose 2B from citrated platelet-rich plasma. Monocytes of 6 SLE patients and 3 healthy donors were isolated using density gradient centrifugation of whole citrated blood followed by negative immune-separation. The >96% purity of the isolated monocytes was confirmed using flow cytometry and the viability of the isolated monocytes assessed by exclusion of trypan blue was 95-99%. The ultrastructure of platelets and monocytes from SLE patients and healthy donors was studied with transmission electron microscopy. A total 192 individual platelets and 154 monocytes from SLE patients were analyzed and compared with 60 individual platelets and 22 monocytes from healthy donors.Results . Most platelets from SLE patients and healthy donors had a normal discoid shape without or with 1-2 short filopodia. Intracellular components (α-granules, dense granules, open canalicular system, lysosomes and mitochondria) were well visualized, albeit somewhat less clear in the SLE platelets. The biggest morphological difference between platelets from healthy subjects and SLE patients was revealed in the platelet plasma membrane. All platelets from healthy donors had normal ultrastructure of the plasma membrane, while the membrane of SLE platelets was very often (in 70% of the cells) shaggy and rough and not as smooth as in normal platelets. About 38% of the SLE platelets were fully bordered with a shaggy plasma membrane, while 32% of the platelets had only partially abnormal plasma membrane. Cells from a patient with anti-cardiolipin and anti-beta-2-glypoprotein antibodies had the most striking membrane abnormalities that were found in 100% of platelets analyzed.Monocytes with normal ultrastructure (ruffled cell surface, formation of filopodia, an indented nucleus, and uniform chromatin distribution) comprised 64% of the total in the SLE patients and 95% in the healthy donors. Unlike monocytes from healthy subjects, 19% of monocytes from the SLE patients had ultrastructural signs of apoptosis with a smooth cell surface, electron-dense cytoplasm, mitochondrial swelling, and a spherical nucleus with perinuclear accumulation of condensed chromatin or condensation of chromatin into dense spherical particles. About 17% of monocytes from the SLE patients and 5% from the healthy donors had necrotic features, such as smooth cell surface without or with quite a few filopodia, low electron-density cytoplasm, small electron-dense mitochondria, and spherical pyknotic nucleus with more diffuse euchromatin. The SLE patients with a high titer of anti-dsDNA antibodies (>200 IU/ml) had significantly more apoptotic (25%, p<0.01, χ2-test) and necrotic (20%, p<0.05) monocytes than those of the SLE patients with a normal titer of anti-dsDNA antibodies (<25 IU/ml) (6% and 10%, respectively).Conclusions . In SLE patients, most platelets have abnormal ultrastructure of the plasma membrane (loss of smoothness, shagginess), likely due to deposition of pathological immune complexes. A substantial fraction of monocytes in SLE undergo morphological alternations of apoptotic or necrotic types. Such alterations were more pronounced in patients with an increased level of anti-cardiolipin, anti-beta-2-glypoprotein, and anti-dsDNA autoantibodies. The results confirm the involvement of both platelets and monocytes in the pathogenesis of SLE, including functional and structural alterations associated with increased production of various autoantibodies.The work was supported by the Program for Competitive Growth at Kazan Federal University. DisclosuresNo relevant conflicts of interest to declare.

Delayed presentation of inflammatory breast carcinoma during the COVID-19 pandemic.

BackgroundBreast cancer may present with distinct cutaneous manifestations that may be paraneoplastic or secondary to direct skin infiltration, distant skin metastases, or dermal lymphatic tumor embolization (inflammatory breast carcinoma).Case reportA 51-year-old Asian woman visited the emergency care department during the outbreak of COVID-19 in Northern Italy. About 6 months before, she had noted the onset of right breast swelling accompanied by skin redness and itching. She never consulted a physician, and, over time, the local skin condition progressed to a large scaly plaque covering the entire breast surface including the nipple. At presentation, abduction of the right upper limb was impaired due to severe shoulder pain. CT scan showed the presence of bilateral breast masses with necrotic and colliquative features, and multiple skeletal, nodal, pulmonary, and brain images suggestive of metastases. An ultrasound-guided core biopsy of the contralateral breast showed grade 2 non-special type infiltrating carcinoma. The patient was referred to the breast oncology unit and is currently being treated with aromatase inhibitors and chemotherapy.ConclusionThe COVID-19 pandemic has disrupted the entire spectrum of oncological care including breast cancer. Hopefully, telemedicine will contribute to increase patients’ confidence and will provide earlier diagnosis and treatment while minimizing the risk of contagion.

Sodium iodate induces ferroptosis in human retinal pigment epithelium ARPE-19 cells

Sodium iodate (SI) is a widely used oxidant for generating retinal degeneration models by inducing the death of retinal pigment epithelium (RPE) cells. However, the mechanism of RPE cell death induced by SI remains unclear. In this study, we investigated the necrotic features of cultured human retinal pigment epithelium (ARPE-19) cells treated with SI and found that apoptosis or necroptosis was not the major death pathway. Instead, the death process was accompanied by significant elevation of intracellular labile iron level, ROS, and lipid peroxides which recapitulated the key features of ferroptosis. Ferroptosis inhibitors deferoxamine mesylate (DFO) and ferrostatin-1(Fer-1) partially prevented SI-induced cell death. Further studies revealed that SI treatment did not alter GPX4 (glutathione peroxidase 4) expression, but led to the depletion of reduced thiol groups, mainly intracellular GSH (reduced glutathione) and cysteine. The study on iron trafficking demonstrated that iron influx was not altered by SI treatment but iron efflux increased, indicating that the increase in labile iron was likely due to the release of sequestered iron. This hypothesis was verified by showing that SI directly promoted the release of labile iron from a cell-free lysate. We propose that SI depletes GSH, increases ROS, releases labile iron, and boosts lipid damage, which in turn results in ferroptosis in ARPE-19 cells.

Short-term whole body cigarette smoke exposure induces regional differences in cellular response in the mouse larynx.

The larynx is an essential organ in the respiratory tract and necessary for airway protection, respiration, and phonation. Cigarette smoking is a significant risk factor associated with benign and malignant laryngeal diseases. Despite this association, the underlying mechanisms by which cigarette smoke (CS) drives disease development are not well elucidated. In the current study, we developed a short-term murine whole body inhalation model to evaluate the first CS-induced cellular responses in the glottic [i.e. vocal fold (VF)] and subglottic regions of the larynx. Specifically, we investigated epithelial cell proliferation, cell death, surface topography, and mucus production, at various time points (1 day, 5 days, 10 days) after ∼ 2 h exposure to 3R4F cigarettes (Delivered dose: 5.6968 mg/kg per cigarette) and following cessation for 5 days after a 5 day CS exposure (CSE). CSE elevated levels of BrdU labeled proliferative cells and p63 labeled epithelial basal cells on day 1 in the VF. CSE increased proliferative cells in the subglottis at days 5, 10 and following cessation in the subglottis. Cleaved caspase-3 apoptotic activity was absent in VF at all time points and increased at day 1 in the subglottis. Evaluation of the VF surface by scanning electron microscopy (SEM) revealed significant epithelial microprojection damage at day 10 and early signs of necrosis at days 5 and 10 post-CSE. SEM visualizations additionally indicated the presence of deformed cilia at days 5 and 10 after CSE and post-cessation in the respiratory epithelium lined subglottis. In terms of mucin content, the impact of short-term CSE was observed only at day 10, with decreasing acidic mucin levels and increasing neutral mucin levels. Overall, these findings reveal regional differences in murine laryngeal cellular responses following short-term CSE and provide insight into potential mechanisms underlying CS-induced laryngeal disease development.

Cytomorphological profile of lymph nodes at a tertiary care centre in Uttar Pradesh: A retrospective study

Fine Needle Aspiration Cytology (FNAC) is a quick, reliable, minimally invasive and cost effective diagnostic modality for evaluation of superficial lymph nodes and has been accepted as a first line investigation in most centres. The aim of the study was to identify the patterns of pathologies in aspiration cytology of lymph nodes in a tertiary care hospital located in Uttar Pradesh, India. The study was undertaken as a retrospective study using data in the Department of Pathology, G.S Medical College and Hospital, Uttar Pradesh, India. 235 lymph node FNACs that were performed during the two year span of 2018-19 were analysed. Females comprised 59.6% of the total patients who underwent lymph node FNAC procedure, 95 (40.4%) were males. The youngest patient was a 3 month old and the oldest happened to be 78 years. Lymph node enlargement was traced to malignant processes exclusively above 40 years of age. Uptill that age, tuberculosis seems to be the major cause underlying lymphadenopathy, mostly presenting as granulomatous lymphadenitis. Out of all cases which showed granulomatous or necrotic features, 27% showed the presence of AFB, hence confirming tuberculosis. A vast majority of the FNACs were performed for cervical lymphadenopathy (92.8%). 42.1% of the FNACs revealed reactive morphology. This diagnosis was closely followed by granulomatous lymphadenitis in 29.4% patients. Together with a definite diagnosis of tuberculous lymphadenitis, patients with granulomatous lymphadenitis comprised 42.6% of all diagnoses. 12 patients received a diagnosis of suppurative lymphadenitis, 9 received necrotizing lymphadenitis. 12 patients were diagnosed as metastatic carcinoma and 1 was suggestive of lymphoma. Most of the lymph nodes whose sizes were under 1 cm were reactive on cytology. Lymphadenopathy due to malignant etiology presented with lymph nodes measuring at least 2 cm. Granulomatous/tuberculous lymphadenitis presented in lymph nodes of varying sizes, from as small as 0.5 cm to larger ones.