Effects of Modified Glucosamine on the Chondrogenic Potential of Circulating Stem Cells under Experimental Inflammation.

Glucosamine (GlcN) is a glycosaminoglycan (GAGs) constituent in connective tissues. It is naturally produced by our body or consumed from diets. In the last decade, in vitro and in vivo trials have demonstrated that the administration of GlcN or its derivates has a protective effect on cartilage when the balance between catabolic and anabolic processes is disrupted and cells are no longer able to fully compensate for the loss of collagen and proteoglycans. To date, these benefits are still controversial because the mechanism of action of GlcN is not yet well clarified. In this study, we have characterized the biological activities of an amino acid (AA) derivate of GlcN, called DCF001, in the growth and chondrogenic induction of circulating multipotent stem cells (CMCs) after priming with tumor necrosis factor-alpha (TNFα), a pleiotropic cytokine commonly expressed in chronic inflammatory joint diseases. In the present work, stem cells were isolated from the human peripheral blood of healthy donors. After priming with TNFα (10 ng/mL) for 3 h, cultures were treated for 24 h with DCF001 (1 µg/mL) dissolved in a proliferative (PM) or chondrogenic (CM) medium. Cell proliferation was analyzed using a Corning® Cell Counter and trypan blue exclusion technique. To evaluate the potentialities of DCF001 in counteracting the inflammatory response to TNFα, we measured the amount of extracellular ATP (eATP) and the expression of adenosine-generating enzymes CD39/CD73, TNFα receptors, and NF-κB inhibitor IκBα using flow cytometry. Finally, total RNA was extracted to perform a gene expression study of some chondrogenic differentiation markers (COL2A1, RUNX2, and MMP13). Our analysis has shed light on the ability of DCF001 to (a) regulate the expression of CD39, CD73, and TNF receptors; (b) modulate eATP under differentiative induction; (c) enhance the inhibitory activity of IκBα, reducing its phosphorylation after TNFα stimulation; and (d) preserve the chondrogenic potentialities of stem cells. Although preliminary, these results suggest that DCF001 could be a valuable supplement for ameliorating the outcome of cartilage repair interventions, enhancing the efficacy of endogenous stem cells under inflammatory stimuli.

The kinesin Eg5 inhibitor K858 exerts antiproliferative and proapoptotic effects and attenuates the invasive potential of head and neck squamous carcinoma cells.

Our group recently demonstrated that K858, an inhibitor of motor kinesin Eg5, has important antiproliferative and apoptotic effects on breast cancer, prostatic cancer, melanoma and glioblastoma cells. Since high levels of kinesin Eg5 expression have been correlated with a poor prognosis in laryngeal carcinoma, we decided to test the anticancer activity of K858 toward this tumor, which belongs to the group of head and neck squamous cell carcinomas (HNSCCs). These cancers are characterized by low responsiveness to therapy. The effects of K858 on the proliferation and assembly of mitotic spindles of three human HNSCC cell lines were studied using cytotoxicity assays and immunofluorescence for tubulin. The effect of K858 on the cell cycle was analyzed by FACS. The expression levels of cyclin B1 and several markers of apoptosis and invasion were studied by Western blot. Finally, the negative regulation of the malignant phenotype by K858 was evaluated by an invasion assay. K858 inhibited cell replication by rendering cells incapable of developing normal bipolar mitotic spindles. At the same time, K858 blocked the cell cycle in the G2 phase and induced the accumulation of cytoplasmic cyclin B and, eventually, apoptosis. Additionally, K858 inhibited cell migration and attenuated the malignant phenotype. The data described confirm that kinesin Eg5 is an interesting target for new anticancer strategies and suggest that this compound may be a powerful tool for an alternative therapeutic approach to HNSCCs.

Biochemical Functions and Clinical Characterizations of the Sirtuins in Diabetes-Induced Retinal Pathologies.

Diabetic retinopathy (DR) is undoubtedly one of the most prominent causes of blindness worldwide. This pathology is the most frequent microvascular complication arising from diabetes, and its incidence is increasing at a constant pace. To date, the insurgence of DR is thought to be the consequence of the intricate complex of relations connecting inflammation, the generation of free oxygen species, and the consequent oxidative stress determined by protracted hyperglycemia. The sirtuin (SIRT) family comprises 7 histone and non-histone protein deacetylases and mono (ADP-ribosyl) transferases regulating different processes, including metabolism, senescence, DNA maintenance, and cell cycle regulation. These enzymes are involved in the development of various diseases such as neurodegeneration, cardiovascular pathologies, metabolic disorders, and cancer. SIRT1, 3, 5, and 6 are key enzymes in DR since they modulate glucose metabolism, insulin sensitivity, and inflammation. Currently, indirect and direct activators of SIRTs (such as antagomir, glycyrrhizin, and resveratrol) are being developed to modulate the inflammation response arising during DR. In this review, we aim to illustrate the most important inflammatory and metabolic pathways connecting SIRT activity to DR, and to describe the most relevant SIRT activators that might be proposed as new therapeutics to treat DR.

Understanding Drivers of Ocular Fibrosis: Current and Future Therapeutic Perspectives.

Ocular fibrosis leads to severe visual impairment and blindness worldwide, being a major area of unmet need in ophthalmology and medicine. To date, the only available treatments are antimetabolite drugs that have significant potentially blinding side effects, such as tissue damage and infection. There is thus an urgent need to identify novel targets to prevent/treat scarring and postsurgical fibrosis in the eye. In this review, the latest progress in biological mechanisms underlying ocular fibrosis are discussed. We also summarize the current knowledge on preclinical studies based on viral and non-viral gene therapy, as well as chemical inhibitors, for targeting TGFß or downstream effectors in fibrotic disorders of the eye. Moreover, the role of angiogenetic and biomechanical factors in ocular fibrosis is discussed, focusing on related preclinical treatment approaches. Moreover, we describe available evidence on clinical studies investigating the use of therapies targeting TGFß-dependent pathways, angiogenetic factors, and biomechanical factors, alone or in combination with other strategies, in ocular tissue fibrosis. Finally, the recent progress in cell-based therapies for treating fibrotic eye disorders is discussed. The increasing knowledge of these disorders in the eye and the promising results from testing of novel targeted therapies could offer viable perspectives for translation into clinical use.

Design, Synthesis and Biological Evaluation of New Pyrimidine Derivatives as Anticancer Agents.

BACKGROUND: Anticancer drug resistance is a challenging phenomenon of growing concern which arises from alteration in drug targets. Despite the fast speed of new chemotherapeutic agent design, the increasing prevalence of this phenomenon requires further research and treatment development. Recently, we reported a new aminopyrimidine compound-namely RDS 344-as a potential innovative anticancer agent. METHODS: Herein, we report the design, synthesis, and anti-proliferative activity of new aminopyrimidine derivatives structurally related to RDS 3442 obtained by carrying out substitutions at position 6 of the pyrimidine core and/or on the 2-aniline ring of our hit. The ability to inhibit cell proliferation was evaluated on different types of tumors, glioblastoma, triple-negative breast cancer, oral squamous cell carcinomas and colon cancer plus on human dermal fibroblasts chosen as control of normal cells. RESULTS: The most interesting compound was the N-benzyl counterpart of RDS 3442, namely 2a, that induced a significant decrease in cell viability in all the tested tumor cell lines, with EC50s ranging from 4 and 8 µM, 4-13 times more active of hit. CONCLUSIONS: These data suggest a potential role for this class of molecules as promising tool for new approaches in treating cancers of different histotype.

Anatomical variations of the pyramidalis muscle: a systematic review and meta-analysis.

PURPOSE: To provide a comprehensive evidence-based assessment of the anatomical characteristics of the pyramidalis muscle (PM). MATERIALS AND METHODS: A thorough systematic search of the literature through August 31st 2020 was conducted on major electronic databases PubMed, Scopus and Web of Science (WOS) to identify studies eligible for inclusion. Data were extracted and pooled into a meta-analysis using MetaFor package in R and MetaXL. A random-effects model was applied. The primary outcome of interest was the prevalence of PM. The secondary outcomes were the dimensions (length and width) of the PM. RESULTS: A total of 11 studies (n = 787 patients; 1548 sides) were included in the meta-analysis. The multinomial pooled prevalence estimate (PPE) for a bilateral absence of the PM was 11.3% (95% CI [7.2%, 16.2%], 82.3% (95% CI [76.2%, 87.6%]) for a bilateral presence, and 6.3% (95% CI [3.3%, 10.2%]) for a unilateral presence. Of four studies (n = 37 patients) that reported the side of a unilateral presence, the PPE of a unilateral right-side presence was 42.2% (95% CI [23.0%, 62.3%]) compared to 57.8% for a unilateral left-side presence (95% CI [37.7%, 77.0%]). The mean length of the PM displayed high levels of heterogeneity, ranging from 3.12 to 12.50 cm. CONCLUSION: The pyramidalis muscle is a rather constant anatomical structure being present in approximately 90% of individuals.

Discovery of a pyrimidine compound endowed with antitumor activity.

Recently, some synthetic nitrogen-based heterocyclic molecules, such as PJ34, have shown pronounced antitumor activity. Therefore, we designed and synthesized new derivatives characterized by a nitrogen-containing scaffold and evaluated their antiproliferative properties in tumor cells. We herein report the effects of three newly synthesized compounds on cell lines from three different human cancers: triple-negative breast cancer, colon carcinoma and glioblastoma. We found that two of these compounds did not affect proliferation, while the third significantly inhibited replication of the three cell lines. Moreover, this third molecule at 20 µM led to the upregulation of p21 and p27 and blockage of the cell cycle at G0/G1; in addition, it induced apoptosis in all three cell lines when used at higher concentrations (30-50 µM). The results demonstrate that this compound is a potent inhibitor of replication, an inducer of apoptosis and a negative regulator of cell cycle progression for cancer cells of different histotypes. Our data suggest a potential role for this new molecule as an interesting and powerful tool for new approaches in treating various cancers.

Inequity in palliative care service full utilisation among patients with advanced cancer: a retrospective Cohort study.

Background: Advanced cancer patients often die in hospital after receiving needless, aggressive treatment. Although palliative care improves symptom management, barriers to accessing palliative care services affect its utilisation, and such disparities challenge the equitable provision of palliative care. This study aimed to identify which factors are associated with inequitable palliative care service utilisation among advanced cancer patients by applying the Andersen Behavioural Model of Health Services Use.Material and methods: This was a retrospective cohort study using administrative healthcare data. A total of 13,656 patients residing in the Lazio region of Italy, who died of an advanced cancer-related cause-either in hospital or in a specialised palliative care facility-during the period of 2012-2016 were included in the study. Potential predictors of specialised palliative service utilisation were explored by grouping the following factors: predisposing factors (i.e., individuals' characteristics), enabling factors (i.e., systemic/structural factors) and need factors (i.e., type/severity of illness).Results: The logistic hierarchical regression showed that older patients (odds ratio [OR] = 1.45; <0.0001) of Caucasian ethnicity (OR = 4.17; 0.02), with a solid tumour (OR = 1.87; <0.0001) and with a longer survival time (OR = 2.09; <0.0001) were more likely to be enrolled in a palliative care service. Patients who lived farther from a specialised palliative care facility (OR = 0.13; <0.0001) and in an urban area (OR = 0.58; <0.0001) were less likely to be enrolled.Conclusion: This study found that socio-demographic (age, ethnicity), clinical (type of tumour, survival time) and organisational (area of residence, distance from service) factors affect the utilisation of specialised palliative care services. The fact that service utilisation is not only a function of patients' needs but also of other aspects demonstrates the presence of inequity in access to palliative care among advanced cancer patients.

Osteopontin in vitreous and idiopathic epiretinal membranes.

PURPOSE: To investigate osteopontin (OPN) expression in vitreous and in related idiopathic epiretinal membranes (ERMs), with respect to VEGF-A, IL8, MIP1α, IL6, and IL33, and correlate OPN expression with disease staging. METHODS: Fifteen (15) vitreous and allied ERMs were collected at the time of therapeutic vitreoretinal surgery. Additional 5 vitreous and 10 ERMs (historical collection) were used. Biochemical and molecular analysis of OPN was performed in clear vitreous, vitreal pelleted cells, and ERMs. Double-immunofluorescence analysis (OPN - GFAP and OPN - αSMA) was performed on paraffin and whole-mounted ERMs. Vitreal OPN levels were correlated to those of VEGF-A, IL8, MIP1α, IL6, and IL33. RESULTS: High OPN levels were observed in vitreal samples, and OPN transcripts were amplified in vitreal cells and related ERMs. OPN immunoreactivity was found in ERMs, mainly in GFAP-bearing (Muller cells) and to a less extend in αSMA-expressing (myofibroblasts) cells. OPN levels were highest at early stages of ERM formation and positively correlated to VEGF-A and MIP1α. CONCLUSIONS: High OPN levels in vitreous, OPN transcripts in vitreal cells/ERMs, OPN immunoreactivity in activated Müller cells and contractile myofibroblasts, as well as the correlation with VEGF-A and MIP1α fulfill the potential involvement of OPN in both inflammation and tissue remodeling that takes part in vitreoretinal interface disorders. The highest OPN levels at early stages of ERM formation would prospect OPN as a potential biomarker for disease severity.

Aberrant left hepatic arteries arising from left gastric arteries and their clinical importance.

BACKGROUND: The Aberrant Left Hepatic Artery (ALHA) is replaced when it does not originate from the hepatic artery proper and it is the only supply to that part of the liver, while an accessory artery coexists with a normal artery. The aim of this systematic review is to evaluate the incidence of ALHAs including the one arising from the Left Gastric Artery, also named Hyrtl's artery. METHODS: A literature search in PubMed, SCOPUS, WOS and Google Scholar was performed. The risk of bias was assessed by means of the AQUA tool. The main outcome was the prevalence of ALHA. Secondary outcomes were the prevalence of the accessory and replaced left hepatic arteries. A subgroup analysis was conducted by geographic region and type of evaluation. RESULTS: This review included 57 studies, with a total of 19,284 patients. The majority of the studies involved the use of radiological techniques -especially Angio-CT-and were performed in Asia. The overall risk of bias was moderate. The overall prevalence of the ALHA was 13.52%; the overall prevalence was 8.26% for the Replaced ALHA and 5.55% for the Accessory ALHA. In the 18 studies that employed Michels' classification, Type II had the lowest prevalence (0.36%) and Type VII the highest prevalence (6.62%). DISCUSSION: Some of the studies included did not distinguish between the ''replaced'' and ''accessory'' ALHA (34.25%). Some surgical dissection techniques proved insufficient for the localization of other hepatic arteries. These results suggest that an accurate preoperative radiological evaluation is needed to localize replaced arteries.

Age related changes seen in human cornea in formalin fixed sections and on biomicroscopy in living subjects: A comparison.

The purpose of our experimental research was to assess the effects of aging on the main corneal structures in healthy corneas. Small, human cornea samples were collected from 20 Caucasian subjects during surgery for traumatic lesions to the eye. Ten subjects were adults (mean age 28 years) and 10 were elderly (mean age 76 years). Morphological analysis was carried out using light microscopy and electron microscopy. Another 40 patients (20 young: mean age < 30 years; 20 elderly: mean age > 70 years) were studied in vivo by confocal microscopy. The resulting images were analyzed qualitatively, quantitatively, and statistically. The basic light microscope revealed a decrease in endothelial cell density with age accompanied by an increase in endothelial cell size. Transmission electron microscopy revealed a corneal thinning and a decrease in the number of corneal stromal cells. A marked decrease in stromal nerve fibers was observed in the older subjects compared to the younger ones. Variable pressure scanning electron microscopy (VP-SEM) was used to make surface morphological observations and to determine the chemical composition of in vivo hydrated human corneas. Our results showed the effects of aging on normal corneal morphology highlighting the structural diversity of the corneal layers and revealing an age-related reduction in nerve fibers, thus explaining the decreased corneal sensitivity that may be observed in the elderly. Clin. Anat. 33:245-256, 2020. © 2019 Wiley Periodicals, Inc.

Amyotrophic Lateral Sclerosis: Autoimmune Pathogenic Mechanisms, Clinical Features, and Therapeutic Perspectives.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the progressive death of motor neurons leading to fatal paralysis. The causes of ALS remain unknown; however, evidence supports the presence of autoimmune mechanisms contributing to pathogenesis. Although several environmental factors have been proposed, the only established risk factors are older age, male gender, and a family history of ALS. To date, there are no diagnostic test for ALS, and clinicians rely on the combination of upper motor neuron and lower motor neuron signs in the same body region. The aim of this paper was to provide a comprehensive review of current clinical literature with special focus on the role of autoimmunity in ALS, differential diagnosis, and available therapeutic approaches. Current evidence suggests a contribution of the innate immune system in ALS, with a role of microglial cell activation at the sites of neurodegeneration. The median time from symptom onset to diagnosis of ALS is 14 months, and this time estimate is mainly based on specific clinical signs and exclusion of ALS-like conditions. Several therapeutic approaches have been proposed, including immunosuppressive drugs, to reduce disease progression. Riluzole has been established as the only, although modestly effective, disease modifying therapy, extending mean patient survival by 3to 6 months. Recent advances in understanding the pathophysiology mechanisms of ALS encourage realistic hope for new treatment approaches. To date, the cornerstones of the management of patients with ALS are focused on symptom control, maintaining quality of life and improving survival.

Validity and reliability of the Palliative Care Transition Measure for Caregivers (PCTM-C).

OBJECTIVE: Patients suffering from advanced disease face different care transitions. The transition from acute to palliative care is challenging and may lead to the discontinuity of care. Family caregivers become important sources of information, as patients begin to experience difficulties in coping with emotional transition events. The Care Transition Measure was developed to evaluate care transitions as experienced by the elderly. It has never been used in palliative care. The aim of this study was to test the validity and reliability of a modified version of the Palliative Care Transition Measure, specifically the Palliative Care Transition Measure for Caregivers (PCTM-C). METHOD: The study included two main phases. Phase I focused on the construction of a modified version of the Palliative Care Transition Measure through two focus groups and by computing the content validity index. Phase II focused on testing the psychometric properties of the PCTM-C on 272 family caregivers through confirmatory factor analysis. RESULT: The content validity index for each of the items was higher than 0.80, whereas that for the scale was 0.95. The model tested with confirmatory factor analysis fitted the data well and confirmed that the transition measures referred to communication, integrated care and a trusting-relationship, and therefore the core dimensions of continuity according to existing conceptual models. The internal consistency was high (Cronbach's alpha = 0.94). SIGNIFICANCE OF RESULTS: The PCTM-C proved to be a suitable measure of the quality of such transitions. It may be used in clinical practice as a continuity quality indicator and has the potential to guide interventions to enhance family caregivers' experience of care continuity.

Prevalence, incidence and associated factors of pressure ulcers in home palliative care patients: A retrospective chart review.

BACKGROUND: Terminally ill patients are at high risk of pressure ulcers, which have a negative impact on quality of life. Data about pressure ulcers' prevalence, incidence and associated factors are largely insufficient. AIM: To document the point prevalence at admission and the cumulative incidence of pressure ulcers in terminally ill patients admitted to an Italian home palliative care unit, and to analyse the patients' and caregivers' characteristics associated with their occurrence. DESIGN: Retrospective chart review. SETTING/PARTICIPANTS: Patients ( n = 574) with a life expectancy ⩽6 months admitted to a palliative home care service were included in this study. RESULTS: The prevalence and incidence rates were 13.1% and 13.0%, respectively. The logistic regression models showed body mass index ( p < 0.001), Braden score at risk ( p < 0.001), Karnofsky Performance Scale index <30 ( p < 0.001), patients' female gender, patients' age >70 and >1 caregiver at home as the dichotomous variables predictors of presenting with a pressure ulcer at time of admission and during home palliative care. CONCLUSION: The notable pressure ulcers' incidence and prevalence rates suggest the need to include this issue among the main outcomes to pursue during home palliative care. The accuracy of body mass index, Braden Scale and Karnofsky Performance Scale in predicting the pressure ulcers risk is confirmed. Therefore, they appear as essential tools, in combination with nurses' clinical judgment, for a structured approach to pressure ulcers prevention. Further research is needed to explore the home caregivers' characteristics and attitudes associated with the occurrence of pressure ulcers and the relations between their strategies for pressure ulcer prevention and gender-related patient's needs.

PAR1 activation affects the neurotrophic properties of Schwann cells.

Protease-activated receptor-1 (PAR1) is the prototypic member of a family of four G-protein-coupled receptors that signal in response to extracellular proteases. In the peripheral nervous system, the expression and/or the role of PARs are still poorly investigated. High PAR1 mRNA expression was found in the rat dorsal root ganglia and the signal intensity of PAR1 mRNA increased in response to sciatic nerve transection. In the sciatic nerve, functional PAR1 receptor was reported at the level of non-compacted Schwann cell myelin microvilli of the nodes of Ranvier. Schwann cells are the principal population of glial cells of the peripheral nervous system which myelinate axons playing an important role during axonal regeneration and remyelination. The present study was undertaken in order to determine if the activation of PAR1 affects the neurotrophic properties of Schwann cells. Our results suggest that the stimulation of PAR1 could potentiate the Schwann cell ability to favour nerve regeneration. In fact, the conditioned medium obtained from Schwann cell cultures challenged with a specific PAR1 activating peptide (PAR1 AP) displays increased neuroprotective and neurotrophic properties with respect to the culture medium from untreated Schwann cells. The proteomic analysis of secreted proteins in untreated and PAR1 AP-treated Schwann cells allowed the identification of factors differentially expressed in the two samples. Some of them (such as macrophage migration inhibitory factor, matrix metalloproteinase-2, decorin, syndecan 4, complement C1r subcomponent, angiogenic factor with G patch and FHA domains 1) appear to be transcriptionally regulated after PAR1 AP treatment as shown by RT-PCR.

Lithium limits trimethyltin-induced cytotoxicity and proinflammatory response in microglia without affecting the concurrent autophagy impairment.

Trimethyltin (TMT) is a highly toxic molecule present as an environmental contaminant causing neurodegeneration particularly of the limbic system both in humans and in rodents. We recently described the occurrence of impairment in the late stages of autophagy in TMT-intoxicated astrocytes. Here we show that similarly to astrocytes also in microglia, TMT induces the precocious block of autophagy indicated by the accumulation of the autophagosome marker, microtubule associated protein light chain 3. Consistent with autophagy impairment we observe in TMT-treated microglia the accumulation of p62/SQSTM1, a protein specifically degraded through this pathway. Lithium has been proved effective in limiting neurodegenerations and, in particular, in ameliorating symptoms of TMT intoxication in rodents. In our in vitro model, lithium displays a pro-survival and anti-inflammatory action reducing both cell death and the proinflammatory response of TMT-treated microglia. In particular, lithium exerts these activities without reducing TMT-induced accumulation of light chain 3 protein. In fact, the autophagic block imposed by TMT is unaffected by lithium administration. These results are of interest as defects in the execution of autophagy are frequently observed in neurodegenerative diseases and lithium is considered a promising therapeutic agent for these pathologies. Thus, it is relevant that this cation can still maintain its pro-survival and anti-inflammatory role in conditions of autophagy block. Copyright © 2016 John Wiley & Sons, Ltd.

Role of parnaparin in atherosclerosis.

Atherosclerosis is characterized by a proliferation of vascular smooth muscle cells (VSMCs) and their migration to the intima, which induces thickening of the intima itself, but the mechanism remains poorly understood. Low molecular weight heparin (LMWH) inhibits the proliferation of VSMCs. Previous studies have shown that a LMWH, parnaparin (PNP), acts on the processes of atherogenesis and atheroprogression in experimental animal models. The aim of this study was to investigate the involvement of oxidative stress, inflammation and VSMCs in the regulation of vascular wall homeostasis. We also considered the possibility of restoring vascular pathological changes using PNP treatment. In order to evaluate vascular remodelling in this study we have analysed the morphological changes in aortas of an animal model of atherosclerosis, apolipoprotein E-deficient mice (ApoE-/-) fed with a normal or a western diet without treatment or treated with PNP. We also analysed, by immunohistochemistry, the expression of proteins linked to atherogenesis and atheroprogression - an enzyme involved in oxidative stress, iNOS, examples of inflammatory mediators, such as tumour necrosis factor alpha (TNF-α), interleukins 1 and 6 (IL-1 and IL-6), and markers of VSMC changes, in particular plasminogen activator inhibitor-1 and thrombospondin-1 (PAI-1 and TSP-1). Our results could suggest that PNP downregulates VSMC proliferation and migration, mediated by PAI-1 and TSP-1, and reduces inflammation and oxidative stress in vessels. These data suggested that LMWH, in particular PNP, could be a theoretically practical tool in the prevention of atherosclerotic vascular modification.

Milestones and recent discoveries on cell death mediated by mitochondria and their interactions with biologically active amines.

Mitochondria represent cell "powerhouses," being involved in energy transduction from the electrochemical gradient to ATP synthesis. The morphology of their cell types may change, according to various metabolic processes or osmotic pressure. A new morphology of the inner membrane and mitochondrial cristae, significantly different from the previous one, has been proposed for the inner membrane and mitochondrial cristae, based on the technique of electron tomography. Mitochondrial Ca(2+) transport (the transporter has been isolated) generates reactive oxygen species and induces the mitochondrial permeability transition of both inner and outer mitochondrial membranes, leading to induction of necrosis and apoptosis. In the mitochondria of several cell types (liver, kidney, and heart), mitochondrial oxidative stress is an essential step in the induction of cell death, although not in brain, in which the phenomenon is caused by a different mechanism. Mitochondrial permeability transition drives both apoptosis and necrosis, whereas mitochondrial outer membrane permeability is characteristic of apoptosis. Adenine nucleotide translocase remains the most important component involved in membrane permeability, with the opening of the transition pore, although other proteins, such as ATP synthase or phosphate carriers, have been proposed. Intrinsic cell death is triggered by the release from mitochondria of proteic factors, such as cytochrome c, apoptosis inducing factor, and Smac/DIABLO, with the activation of caspases upon mitochondrial permeability transition or mitochondrial outer membrane permeability induction. Mitochondrial permeability transition induces the permeability of the inner membrane in sites in contact with the outer membrane; mitochondrial outer membrane permeability forms channels on the outer membrane by means of various stimuli involving Bcl-2 family proteins. The biologically active amines, spermine, and agmatine, have specific functions on mitochondria which distinguish them from other amines. Enzymatic oxidative deamination of spermine by amine oxidases in tumor cells may produce reactive oxygen species, leading to transition pore opening and apoptosis. This process could be exploited as a new therapeutic strategy to combat cancer.

Age and diabetes related changes of the retinal capillaries: An ultrastructural and immunohistochemical study.

Normal human aging and diabetes are associated with a gradual decrease of cerebral flow in the brain with changes in vascular architecture. Thickening of the capillary basement membrane and microvascular fibrosis are evident in the central nervous system of elderly and diabetic patients. Current findings assign a primary role to endothelial dysfunction as a cause of basement membrane (BM) thickening, while retinal alterations are considered to be a secondary cause of either ischemia or exudation. The aim of this study was to reveal any initial retinal alterations and variations in the BM of retinal capillaries during diabetes and aging as compared to healthy controls. Moreover, we investigated the potential role of vascular endothelial growth factor (VEGF) and pro-inflammatory cytokines in diabetic retina.Transmission electron microscopy (TEM) was performed on 46 enucleated human eyes with particular attention to alterations of the retinal capillary wall and Müller glial cells. Inflammatory cytokines expression in the retina was investigated by immunohistochemistry.Our electron microscopy findings demonstrated that thickening of the BM begins primarily at the level of the glial side of the retina during aging and diabetes. The Müller cells showed numerous cytoplasmic endosomes and highly electron-dense lysosomes which surrounded the retinal capillaries. Our study is the first to present morphological evidence that Müller cells start to deposit excessive BM material in retinal capillaries during aging and diabetes. Our results confirm the induction of pro-inflammatory cytokines TNF-α and IL-1ß within the retina as a result of diabetes.These observations strongly suggest that inflammatory cytokines and changes in the metabolism of Müller glial cells rather than changes in of endothelial cells may play a primary role in the alteration of retinal capillaries BM during aging and diabetes.