Nonspecific Orbital Inflammation (NSOI): Unraveling the Molecular Pathogenesis, Diagnostic Modalities, and Therapeutic Interventions.

Nonspecific orbital inflammation (NSOI), colloquially known as orbital pseudotumor, sometimes presents a diagnostic and therapeutic challenge in ophthalmology. This review aims to dissect NSOI through a molecular lens, offering a comprehensive overview of its pathogenesis, clinical presentation, diagnostic methods, and management strategies. The article delves into the underpinnings of NSOI, examining immunological and environmental factors alongside intricate molecular mechanisms involving signaling pathways, cytokines, and mediators. Special emphasis is placed on emerging molecular discoveries and approaches, highlighting the significance of understanding molecular mechanisms in NSOI for the development of novel diagnostic and therapeutic tools. Various diagnostic modalities are scrutinized for their utility and limitations. Therapeutic interventions encompass medical treatments with corticosteroids and immunomodulatory agents, all discussed in light of current molecular understanding. More importantly, this review offers a novel molecular perspective on NSOI, dissecting its pathogenesis and management with an emphasis on the latest molecular discoveries. It introduces an integrated approach combining advanced molecular diagnostics with current clinical assessments and explores emerging targeted therapies. By synthesizing these facets, the review aims to inform clinicians and researchers alike, paving the way for molecularly informed, precision-based strategies for managing NSOI.

An Overview of the Dry Eye Disease in Sjögren's Syndrome Using Our Current Molecular Understanding.

Sjögren's syndrome is a chronic and insidious auto-immune disease characterized by lymphocyte infiltration of exocrine glands. The patients typically present with ocular surface diseases related to dry eye and other systemic manifestations. However, due to the high prevalence of dry eye disease and the lack of objective and clinically reliable diagnostic tools, discriminating Sjögren's syndrome dry eye (SSDE) from non-Sjögren's syndrome dry eye (NSSDE) remains a challenge for clinicians. Diagnosing SS is important to improve the quality of life of patients through timely referral for systemic workups, as SS is associated with serious systemic complications such as lymphoma and other autoimmune diseases. The purpose of this article is to describe the current molecular understanding of Sjögren's syndrome and its implications for novel diagnostic modalities on the horizon. A literature review of the pre-clinical and clinical studies published between 2016 and 2022 was conducted. The SSDE pathophysiology and immunology pathways have become better understood in recent years. Novel diagnostic modalities, such as tear and saliva proteomics as well as exosomal biomarkers, provide hope on the horizon.

Energetic metabolic reprogramming in Jurkat DFF40-deficient cancer cells.

DNA fragmentation factor 40 (DFF40), or the caspase-activated DNase (CAD), is an endonuclease specific for double-stranded DNA. Alterations in its function and expression have been linked to apoptosis resistance, a mechanism likely used by cancer cells. However, how the DFF40-related apoptosis resistance pathway occurs remains unclear. Here, we sought to determine if DFF40 expression could be linked to cell metabolism through the regulation of mitochondrial integrity and function. We demonstrated that DFF40-deficient cells are more resistant to staurosporine and tributyltin (TBT)-induced apoptosis, and express higher levels of Mcl-1 at basal state. Treatment with TBT induces higher Bcl-2 and caspase-9 mRNA transcripts in DFF40 KO Jurkat cells, as well as enhanced Bcl-2 phosphorylation. A loss of DFF40 expression induces a higher mitochondrial mass, mtDNA copy number, mitochondrial membrane potential, and glycolysis rates in resting T cells. DFF40-deficient cells exhibit the Warburg effect phenotype, where they rely significantly more on glycolysis than oxidative phosphorylation and have a higher proliferative state, demonstrated by a higher Ki-67 transcription factor expression and AKT phosphorylation. Finally, we demonstrated with cell fractioning that DFF40 can translocate to the mitochondria following apoptosis induction. Our study reveals that DFF40 may act as a regulator of mitochondria during cell death and its loss could compromise mitochondrial integrity and cause an energetic reprogramming in pathologies such as cancer.

Review of cancer cell resistance mechanisms to apoptosis and actual targeted therapies.

The apoptosis pathway is a programmed cell death mechanism that is crucial for cellular and tissue homeostasis and organ development. There are three major caspase-dependent pathways of apoptosis that ultimately lead to DNA fragmentation. Cancerous cells are known to highly regulate the apoptotic pathway and its role in cancer hallmark acquisition has been discussed over the past decades. Numerous mutations in cancer cell types have been reported to be implicated in chemoresistance and treatment outcome. In this review, we summarize the mutations of the caspase-dependant apoptotic pathways that are the source of cancer development and the targeted therapies currently available or in trial.

DFF40 deficiency in cancerous T cells is implicated in chemotherapy drug sensitivity and resistance through the regulation of the apoptotic pathway.

The regulation of the apoptotic pathway is one of the most studied mechanisms regarding cancer cell resistance. Many mutations have been linked to drug resistance. The DNA fragmentation factor 40 (DFF40) has been gaining interest regarding cancer cell response to chemotherapy and patient outcomes. Glioblastomas and uterine leiomyosarcomas have been shown to have a downregulation of DFF40 expression, conferring a poor patient prognosis. In concordance with these observations, in this study, we showed that DFF40 gene is also downregulated in breast, endocervical, ovarian, lung, pancreas and glioblastomas. DFF40 is the endonuclease responsible of DNA fragmentation during apoptosis. In this study, we sought to determine if a DFF40 deficiency in Jurkat T cells could impact the sensitivity to conventional chemotherapy drugs. CRISPR-cas9 generated DFF40 knockout (DFF40 KO) stable Jurkat cells and wild-type (DFF40 WT) cells were treated with different antimetabolites and topoisomerase II (TOP2) inhibitors, and cell viability was subsequently assessed. DFF40 deficient cells show chemoresistance to antimetabolites (e.g. methotrexate, 6-mercaptopurine and cytarabine) and surprisingly, they are more sensitive to TOP2 inhibitors (e.g. etoposide and teniposide). DFF40 deficient cells exposed to cytarabine present lower phosphatidylserine translocation levels to the outer cell membrane layer. Etoposide exposure in DFF40 deficient cells induces higher mortality levels and downregulation of Bcl-xL cells compared to DFF40 expressing T cells. The abolition of DFF40 expression in Jurkat cells significantly impairs histone H2AX phosphorylation following etoposide and cytarabine treatments. Our findings suggest that DFF40 is a novel key target in cancer cell resistance that potentially regulates genomic stability.

The RyfA small RNA regulates oxidative and osmotic stress responses and virulence in uropathogenic Escherichia coli.

Urinary tract infections (UTIs) are a common bacterial infectious disease in humans, and strains of uropathogenic Escherichia coli (UPEC) are the most frequent cause of UTIs. During infection, UPEC must cope with a variety of stressful conditions in the urinary tract. Here, we demonstrate that the small RNA (sRNA) RyfA of UPEC strains is required for resistance to oxidative and osmotic stresses. Transcriptomic analysis of the ryfA mutant showed changes in expression of genes associated with general stress responses, metabolism, biofilm formation and genes coding for cell surface proteins. Inactivation of ryfA in UPEC strain CFT073 decreased urinary tract colonization in mice and the ryfA mutant also had reduced production of type 1 and P fimbriae (pili), adhesins which are known to be important for UTI. Furthermore, loss of ryfA also reduced UPEC survival in human macrophages. Thus, ryfA plays a key regulatory role in UPEC adaptation to stress, which contributes to UTI and survival in macrophages.

The role of the DFF40/CAD endonuclease in genomic stability.

Maintenance of genomic stability in cells is primordial for cellular integrity and protection against tumor progression. Many factors such as ultraviolet light, oxidative stress, exposure to chemical reagents, particularly mutagens and radiation, can alter the integrity of the genome. Thus, human cells are equipped with many mechanisms that prevent these irreversible lesions in the genome, as DNA repair pathways, cell cycle checkpoints, and telomeric function. These mechanisms activate cellular apoptosis to maintain DNA stability. Emerging studies have proposed a new protein in the maintenance of genomic stability: the DNA fragmentation factor (DFF). The DFF40 is an endonuclease responsible of the oligonucleosomal fragmentation of the DNA during apoptosis. The lack of DFF in renal carcinoma cells induces apoptosis without oligonucleosomal fragmentation, which poses a threat to genetic information transfer between cancerous and healthy cells. In this review, we expose the link between the DFF and genomic instability as the source of disease development.

DNA fragmentation factor 40 expression in T cells confers sensibility to tributyltin-induced apoptosis.

DNA fragmentation factor 40 (DFF40), an endonuclease, mediates the final and irreversible step of apoptosis by conducting oligonucleosomal DNA fragmentation. New emerging studies have proposed a role of DFF40 in genomic stability, besides its nuclease activity. Overexpression of DFF40 in tumoral cells increases their sensitivity to chemotherapeutic drugs. In this study, we sought to determine if DFF40 expression influences the toxicity of tributyltin (TBT), a well-known immunotoxic and apoptosis-inducing compound. The strategy used was to knockout DFF40 expression by CRISPR-cas9 method in Jurkat T cells and to determine the toxicity of TBT in DFF40 KO cells and DFF40 WT Jurkat cells. DFF40 KO Jurkat cells show an increase of cell viability following a 24-h TBT exposure (p < 0.05). There is a resistance to TBT-induced apoptosis determined by annexin V/PI am labeling (p < 0.05). Interestingly, the basal level of ROS rises in DFF40 KO Jurkat cells, but ROS production levels after TBT exposure remains at the same basal level. Other apoptosis or DNA damage makers (procaspase-3, caspase-6, and PARP cleavage) are significantly delayed and decreased. DFF40 deficient cells do not present histone H2AX phosphorylation, whereas wild-type cells present a phosphorylation following a 6-h exposure to TBT (p < 0.001). The re-expression of DFF40 in DFF40 KO cells restores the cytotoxic effects of TBT. Overall, these data suggest a role of DFF40 in cells sensitivity to TBT and possibly in DNA stability.