Caspase-8-mediated inflammation but not apoptosis drives death of retinal ganglion cells and loss of visual function in glaucomaa.

Background-: Glaucoma is a complex multifactorial disease where apoptosis and inflammation represent two key pathogenic mechanisms. However, the relative contribution of apoptosis versus inflammation in axon degeneration and death of retinal ganglion cells (RGCs) is not well understood. In glaucoma, caspase-8 is linked to RGC apoptosis, as well as glial activation and neuroinflammation. To uncouple these two pathways and determine the extent to which caspase-8-mediated inflammation and/or apoptosis contributes to the death of RGCs, we used the caspase-8 D387A mutant mouse (Casp8 DA/DA ) in which a point mutation in the auto-cleavage site blocks caspase-8-mediated apoptosis but does not block caspase-8-mediated inflammation. Methods-: Intracameral injection of magnetic microbeads was used to elevate the intraocular pressure (IOP) in wild-type, Fas deficient Faslpr, and Casp8 DA/DA mice. IOP was monitored by rebound tonometry. Two weeks post microbead injection, retinas were collected for microglia activation analysis. Five weeks post microbead injection, visual acuity and RGC function were assessed by optometer reflex (OMR) and pattern electroretinogram (pERG), respectively. Retina and optic nerves were processed for RGC and axon quantification. Two- and five-weeks post microbead injection, expression of the necrosis marker, RIPK3, was assessed by qPCR. Results-: Wild-type, Faslpr, and Casp8 DA/DA mice showed similar IOP elevation as compared to saline controls. A significant reduction in both visual acuity and pERG that correlated with a significant loss of RGCs and axons was observed in wild-type but not in Faslpr mice. The Casp8 DA/DA mice displayed a significant reduction in visual acuity and pERG amplitude and loss of RGCs and axons similar to that in wild-type mice. Immunostaining revealed equal numbers of activated microglia, double positive for P2ry12 and IB4, in the retinas from microbead-injected wild-type and Casp8 DA/DA mutant mice. qPCR analysis revealed no induction of RIPK3 in wild-type or Casp8 DA/DA mice at two- or five-weeks post microbead injection. Conclusions-: Our results demonstrate that caspase-8-mediated extrinsic apoptosis is not involved in the death of RGCs in the microbead-induced mouse model of glaucoma implicating caspase-8-mediated inflammation, but not apoptosis, as the driving force in glaucoma progression. Taken together, these results identify the caspase-8-mediated inflammatory pathway as a potential target for neuroprotection in glaucoma.

Tumor microenvironment signaling and therapeutics in cancer progression.

Tumor development and metastasis are facilitated by the complex interactions between cancer cells and their microenvironment, which comprises stromal cells and extracellular matrix (ECM) components, among other factors. Stromal cells can adopt new phenotypes to promote tumor cell invasion. A deep understanding of the signaling pathways involved in cell-to-cell and cell-to-ECM interactions is needed to design effective intervention strategies that might interrupt these interactions. In this review, we describe the tumor microenvironment (TME) components and associated therapeutics. We discuss the clinical advances in the prevalent and newly discovered signaling pathways in the TME, the immune checkpoints and immunosuppressive chemokines, and currently used inhibitors targeting these pathways. These include both intrinsic and non-autonomous tumor cell signaling pathways in the TME: protein kinase C (PKC) signaling, Notch, and transforming growth factor (TGF-ß) signaling, Endoplasmic Reticulum (ER) stress response, lactate signaling, Metabolic reprogramming, cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) and Siglec signaling pathways. We also discuss the recent advances in Programmed Cell Death Protein 1 (PD-1), Cytotoxic T-Lymphocyte Associated Protein 4 (CTLA4), T-cell immunoglobulin mucin-3 (TIM-3) and Lymphocyte Activating Gene 3 (LAG3) immune checkpoint inhibitors along with the C-C chemokine receptor 4 (CCR4)- C-C class chemokines 22 (CCL22)/ and 17 (CCL17), C-C chemokine receptor type 2 (CCR2)- chemokine (C-C motif) ligand 2 (CCL2), C-C chemokine receptor type 5 (CCR5)- chemokine (C-C motif) ligand 3 (CCL3) chemokine signaling axis in the TME. In addition, this review provides a holistic understanding of the TME as we discuss the three-dimensional and microfluidic models of the TME, which are believed to recapitulate the original characteristics of the patient tumor and hence may be used as a platform to study new mechanisms and screen for various anti-cancer therapies. We further discuss the systemic influences of gut microbiota in TME reprogramming and treatment response. Overall, this review provides a comprehensive analysis of the diverse and most critical signaling pathways in the TME, highlighting the associated newest and critical preclinical and clinical studies along with their underlying biology. We highlight the importance of the most recent technologies of microfluidics and lab-on-chip models for TME research and also present an overview of extrinsic factors, such as the inhabitant human microbiome, which have the potential to modulate TME biology and drug responses.

Ayurvedic formulations amalaki rasayana and rasa sindoor improve age-associated memory deficits in mice by modulating dendritic spine densities.

BACKGROUND: Emerging reports indicate that age-associated cognitive decline begins with the transition from young to middle-aged, and this neurological condition manifests mainly due to the progressive impairment in the adaptive homeostasis process. Moreover, cognitive decline is associated with neurodegenerative changes in older adults. OBJECTIVE: Previous studies have shown that the administration of Ayurvedic formulations restores the homeostatic pathways and ameliorates neurodegeneration in animal models of neurodegenerative diseases. Therefore, we wanted to check whether Ayurvedic formulations can rescue or delay the age-associated cognitive decline in middle-aged mice. MATERIAL AND METHODS: We fed two-month-old mice with amalaki aasayana (AR, 1025 mg/kg per day) or rasa sindoor (RS, 41 mg/kg per day) mixed in a gelatin-based jelly for six months. Mice eating regular chow or blank jelly served as control. Subsequently, we looked at the improvements in the cognitive and behavioural traits of the treated animals. We have also analysed the effect of these formulations on the dendritic processes of neurons, glial activation, and the formation of corpora amylacea. RESULTS: We found a significant improvement in episodic, working- and reference-spatiotemporal memory in animals fed on AR or RS. Microscopic analyses revealed a significant increase in the dendritic spine density in the apical dendrites of the hippocampal pyramidal neurons. The treatment, however, did not significantly affect gliosis and corpora amylacea in the brains. CONCLUSIONS: Both AR and RS showed beneficial effects on memory functions of the middle-aged mice, possibly due to their effect on the dendritic spine densities. Our findings provide strong evidence to conclude that formulations AR and RS can prevent or delay the onset of age-associated cognitive decline.

Age-Dependent Reduction in the Expression Levels of Genes Involved in Progressive Myoclonus Epilepsy Correlates with Increased Neuroinflammation and Seizure Susceptibility in Mouse Models.

Brain aging is characterized by a gradual decline in cellular homeostatic processes, thereby losing the ability to respond to physiological stress. At the anatomical level, the aged brain is characterized by degenerating neurons, proteinaceous plaques and tangles, intracellular deposition of glycogen, and elevated neuroinflammation. Intriguingly, such age-associated changes are also seen in neurodegenerative disorders suggesting that an accelerated aging process could be one of the contributory factors for the disease phenotype. Amongst these, the genetic forms of progressive myoclonus epilepsy (PME), resulting from loss-of-function mutations in genes, manifest symptoms that are common to age-associated disorders, and genes mutated in PME are involved in the cellular homeostatic processes. Intriguingly, the incidence and/or onset of epileptic seizures are known to increase with age, suggesting that physiological changes in the aged brain might contribute to increased susceptibility to seizures. We, therefore, hypothesized that the expression level of genes implicated in PME might decrease with age, thereby leading to a compromised neuronal response towards physiological stress and hence neuroinflammation in the aging brain. Using mice models, we demonstrate here that the expression level of PME genes shows an inverse correlation with age, neuroinflammation, and compromised heat shock response. We further show that the pharmacological suppression of neuroinflammation ameliorates seizure susceptibility in aged animals as well as in animal models for a PME. Taken together, our results indicate a functional role for the PME genes in normal brain aging and that neuroinflammation could be a major contributory player in susceptibility to seizures.

Dexamethasone-induced activation of heat shock response ameliorates seizure susceptibility and neuroinflammation in mouse models of Lafora disease.

Heat shock response (HSR) is a conserved cytoprotective pathway controlled by the master transcriptional regulator, the heat shock factor 1 (HSF1), that activates the expression of heat shock proteins (HSPs). HSPs, as chaperones, play essential roles in minimizing stress-induced damages and restoring proteostasis. Therefore, compromised HSR is thought to contribute to neurodegenerative disorders. Lafora disease (LD) is a fatal form of neurodegenerative disorder characterized by the accumulation of abnormal glycogen as Lafora bodies in neurons and other tissues. The symptoms of LD include progressive myoclonus epilepsy, dementia, and cognitive deficits. LD is caused by the defects in the gene coding laforin phosphatase or the malin ubiquitin ligase. Laforin and malin are known to work upstream of HSF1 and are essential for the activation of HSR. Herein, we show that mice deficient for laforin or malin show reduced levels of HSF1 and their targets in their brain tissues, suggesting compromised HSR; this could contribute to the neuropathology in LD. Intriguingly, treatment of LD animals with dexamethasone, a synthetic glucocorticoid analogue, partially restored the levels of HSF1 and its targets. Dexamethasone treatment was also able to ameliorate the neuroinflammation and susceptibility to induced seizures in the LD animals. However, dexamethasone treatment did not show a significant effect on Lafora bodies or autophagy defects. Taken together, the present study establishes a role for HSR in seizure susceptibility and neuroinflammation and dexamethasone as a potential antiepileptic agent, suitable for further studies in LD.

Trehalose Ameliorates Seizure Susceptibility in Lafora Disease Mouse Models by Suppressing Neuroinflammation and Endoplasmic Reticulum Stress.

Lafora disease (LD) is one of the progressive and fatal forms of a neurodegenerative disorder and is characterized by teenage-onset myoclonic seizures. Neuropathological changes in LD include the formation of abnormal glycogen as Lafora bodies, gliosis, and neuroinflammation. LD is caused by defects in the gene coding for phosphatase (laforin) or ubiquitin ligase (malin). Mouse models of LD, developed by targeted disruption of these two genes, develop most symptoms of LD and show increased susceptibility to induced seizures. Studies on mouse models also suggest that defective autophagy might contribute to LD etiology. In an attempt to understand the specific role of autophagy in LD pathogenesis, in this study, we fed LD animals with trehalose, an inducer of autophagy, for 3 months and looked at its effect on the neuropathology and seizure susceptibility. We demonstrate here that trehalose ameliorates gliosis, neuroinflammation, and endoplasmic reticulum stress and reduces susceptibility to induced seizures in LD animals. However, trehalose did not affect the formation of Lafora bodies, suggesting the epileptic phenotype in LD could be either secondary to or independent of Lafora bodies. Taken together, our results suggest that autophagy inducers can be considered as potential therapeutic molecules for Lafora disease.