Endoplasmic reticulum stress and Nrf2 repression in circulating cells of type 2 diabetic patients without the recommended glycemic goals

Endoplasmic reticulum (ER) stress plays a role in the pathogenesis of type 2 diabetes mellitus (T2DM), with activation of the unfolded protein response (UPR) and ER apoptosis in \u3b2-cells. The aim of the study is investigating the role of the prolonged glycemic, inflammatory, and oxidative impairment as possible UPR and ER apoptosis inductors in triggering the ER stress response and the protective nuclear erythroid-related factor 2 (Nrf2)/antioxidant-related element (ARE) activation in peripheral blood mononuclear cells (PBMC) of T2DM patients without glycemic target. Oxidative stress markers (oxidation product of phospholipid 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine [oxPAPC], and malondialdehyde [MDA]), the UPR and ER apoptosis, the activation of the pro-inflammatory nuclear factor-kappa B (NF-kB) with its inhibitory protein inhibitor-kB\u3b1, and the expression of the protective Nrf2 and heme oxygenase-1 (HO-1) were evaluated in PBMC of 15 T2DM patients and 15 healthy controls (C). OxPAPC concentrations (in PBMC and plasma), MDA levels (in plasma), the expressions of the glucose-regulated protein 78 kDa (or BiP) as representative of UPR, and of the CCAAT/enhancer-binding protein homologous protein as representative of ER apoptosis were significantly higher (p < 0.01) in T2DM with respect to C. IkB\u3b1 expression was significantly lower (p < 0.01) in T2DM as well as Nrf2 and HO-1. In vitro experiments demonstrated that hyperglycemic conditions, if prolonged, were NF-kB inductors, without a corresponding Nrf2/ARE response. In PBMC of T2DM without glycemic target achievement, there is an activation of the UPR and of the ER apoptosis, which may be related to the chronic exposure to hyperglycemia, to the augmented inflammation, and to the augmented oxidative stress, without a corresponding Nrf2/ARE defense activation

A comparison in university students of the amplitude of accommodation determined subjectively

Background: Historically, the push-up and the minus lens methods have been used for the measurements of the amplitude of accommodation, and the differences between the results of these methods are well known. Aim: The purpose of this study was to compare three methods for determining the monocular amplitude of accommodation and consider whether agreement exists between such methods. Setting: The study was conducted at the Optometry Clinic, University of Limpopo. Method: Thirty-four (N = 34) African optometry students participated in this study. There were 20 female and 14 male students. The age range of the participants was 20–34 years. Amplitude of accommodation was measured via the subjective push-up, push-down and minus lens methods only on the right eyes of the sample. Results: The highest average amplitude of accommodation was obtained with the push-up method (10.20 D ± 0.96 D), while the minus lens method produced the smallest mean amplitude of accommodation (9.66 D ± 0.75 D). A higher correlation was found between the push-up and push-down methods (r = 0.80, p = 0.06). The smallest correlation was observed between the push-up and the minus lens methods (r = 0.60, p = 0.062). There were no statistically significant differences between the amplitude of accommodation in male and female students for all three methods (p > 0.005). Conclusion: It seems easier to recognise the point where one can identify a target in pushdown amplitude than the point of first sustained blur in the push-up method. The push-up method tends to overestimate the actual amplitude of accommodation because of the effects of depth of focus. The less evaluated method in the literature is the push-away method; however, further research is necessary to answer the question of which (if any) method is more accurate

SUPERVISED INTENSIVE TRAINING IMPROVES OXIDATIVE DEFENSIVE PATHWAYS AND REDUCES INFLAMMATION IN PERIPHERAL ARTERY DISEASE

Background: recent evidences suggest that exercise training may ameliorate oxidative stress in patients with peripheral arterial disease (PAD) and reduce inflammation. Few data are available on mechanisms involved. Nuclear related factor 2 (Nrf2), regulates the expression of antioxidant enzymes and proteins through the antioxidant response ; Nrf2 is also involved in protection against oxidant stress during the processes of ischemia-reperfusion injury. Also monocytes (CD16+) are recognizesd as an important actor in aterosclerosis. Aim: to evaluate in PAD the effects of controlled exercise training on oxidative stress, Nrf2 gene expression and monocyte subsets. Methods: 24 patients, with PAD at Fontaine stage II, underwent a 3-weeks (5 days per week) period of supervised training with treadmill walking conducted until pain onset with subsequent rest and walking again for total 40 minutes of exercise. At beginning (D0) and at the end of the program (D21) were evaluated: pain-free walking time (PFWT), maximal walking time (MWT), monocyte subsets by flow cytometry, malondialdehyde (MDA), as marker of oxidative stress, total RNA (extracted from PBMCs to evaluate Nrf2 expression by RT-PCR). Results: the 3-week training significantly increased PWFT and MWT (p=0.001); plasma levels of MDA decreased (p=0.002), levels of mRNA Nrf2 increased after the exercise program (p=0.004, figure 1 ); finally, the percentage of inflammatory monocytes decreased, and conversely classical monocytes increased (figure 2). Conclusions: Our study shows that training in PAD patients improves walking time, furthermore it reduces oxidative stress (decrease of MDA) and increases anti-oxidative defensive system enhancing Nrf2-gene expression. The training elicited multiple episodes of ischemia-reperfusion every day and, since Nrf2 is involved in protecting from ischemia-reperfusion injury we can infer that somehow we "trained" also Nrf2 system. The shift of monocytes subtype from an inflammatory to a classic subtype suggests a deep change in inflammation profile of these patients with training

Do oxidized polyunsaturated Fatty acids affect endoplasmic reticulum stress-induced apoptosis in human carotid plaques?

Macrophage apoptosis is involved in atherosclerotic plaque development. The aim of this study was to evaluate the interrelationship between macrophage apoptosis and the endoplasmic reticulum (ER) stress in the tissue around the necrotic core (TANC) and in the periphery (P) of the same carotid plaques derived from patients undergoing carotid endarterectomy. Apoptosis was significantly higher in TANC than in P (p<0.001). mRNA and protein expression of the protein kinase-like ER kinase (Perk) and the nuclear erythroid-related factor 2 (Nrf2)-related survival genes was significantly higher in P than in TANC (p<0.01), while CCAAT/enhancer-binding protein homologous protein (Chop) and the apoptosis-related genes were higher in TANC than in P (p<0.01). The TANC extract was characterized by significantly higher concentrations of oxidized derivatives of polyunsaturated fatty acids (PUFAs) than the P extract (p<0.01). When THP-1 cells were incubated with P or TANC extracts there was a dose-dependent increase of Perk and Nrf2 or of Chop and of the apoptosis-related genes, respectively (p<0.01). Our observations lead to the hypothesis that ER stress induced by oxidized derivatives of PUFAs may promote macrophage apoptosis in TANC and favor the expansion of the necrotic core of the plaques, a major feature responsible for its disruption and acute luminal thrombosis

Il training aerobico intensivo migliora le difese antiossidanti e riduce l'infiammazione in pazienti con arteriopatia degli arti inferiori.

IL TRAINING AEROBICO INTENSIVO MIGLIORA LE DIFESE ANTIOSSIDANTI E RIDUCE L\u2019INFIAMMAZIONE IN PAZIENTI CON ARTERIOPATIA DEGLI ARTI INFERIORI S. De Marchi, U. Garbin*, A. Rigoni, E. Solani*, C. Stranieri *, A. Pasini *, A. Tinelli *, M. Prior , E. Arosio. Sezione di Angiologia, Sezione di Medicina Interna D*, Dipartimento di Medicina, Universit\ue0 di Verona . Introduzione : dati recenti dalla letteratura hanno evidenziato come il training aerobico supervisionato possa ridurre lo stress ossidativo e l\u2019infiammazione in pazienti con arteriopatia obliterante degli arti inferiori (PAD). Tuttavia sono pochi i dati che dimostrino ed esplorino i meccanismi coinvolti in questi favorevoli modifiche. Il Nuclear related factor 2 (Nrf2) \ue8 un importante sistema di trascrizione che regola l\u2019espressione di enzimi antiossidanti e modula la risposta allo stress ossidativo a livello cellulare e del reticolo endoplasmatico (1). Di particolare rilievo risulta il coinvolgimento di Nrf2 nella protezione dallo stresso ossidativo indotto da ischemia-riperfusione (2,3). Sotto il profilo infiammatorio di particolare interesse risulta il ruolo dei monociti (CD16+) che sono riconosciuti come protagonisti nella genesi della placca aterosclerotica. Obiettivo: valutare il pazienti con PAD gli effetti di un ciclo di training fisico controllato sullo stress ossidativo, sull\u2019espressione di Nrf2 e sui sottotipi di monociti. Materiali e Metodi : 24 pazienti, affetti da PAD al II stadio di Fontaine, sono stati sottoposti a 3 settimane (5 giorni per settimana) di training supervisionato con camminata su treadmill effettuata sino alla comparsa di dolore con successiva sosta e ripetizione del cammino per un totale di 40 minuti. All\u2019inizio (D0) ed al termine del programma (D21) i pazienti sono stati valutati per : distanza di marcia libera da dolore (PFWT), distanza massima di marcia (MWT), sottotipi monocitari con citofluorimetria, malondialdeide (MDA) come marcatore di stress ossidativo, RNA totale estratto da PBMCs per valutare l\u2019espressione di Nrf2 attraverso la metodica RT-PCR. Risultati: il training ha incrementato signifcativamente PWFT e MWT (p=0.001), ha ridotto le concentrazioni plasmatiche di MDA (p=0.002 \u2013 figura 1), ha aumentato i livelli di mRNA- Nrf2 (p=0.004, figura 2 ), in fine ha ridotto la percentuale di monociti infiammatori con incremento dei monociti di tipo classico CD14++CD16- vs CD14++CD16+ (figura 3). Conclusioni : il nostro studio ha mostrato che il training nei pazienti PAD migliora l\u2019autonomia di marcia, come noto, ma che al contempo \ue8 in grado di ridurre lo stress ossidativo (decremento di MDA) verosimilmente mediante un potenziamento del sistema di difesa antiossidante stimolato dall\u2019incremento dell\u2019espressione dei geni del sistema Nrf2. Il trianing ha stimolato multipli e subentranti episodi di ischemia-riperfusione di entit\ue0 modesta ma quotidiani, ci\uf2 pu\uf2 aver stimolato Nrf2 essendo questo un meccanismo coinvolto nella protezione dal danno da ischemia-riperfusione. Inoltre , lo shift dei sottotipi monocitari da una forma infiammatoria ad una classica suggerisce un profondo reset del profilo infiammatorio ottenuto in questi pazienti con il training, e dimostrato sul versante cellulare e oltre gli usuali marcatori di fase acuta o le molecole di adesione. In conclusione si potrebbe quasi immaginare che sia stato possibile effettuare un training del sistema Nrf2 oltre che una modifica dell\u2019assetto immunitario. Bibliografia 1. Cominacini L, Mozzini C, Garbin U, Pasini A, Stranieri C, Solani E, Vallerio P, Irene Tinelli A, Fratta Pasini A. Endoplasmic reticulum stress and Nrf2 signaling in cardiovascular diseases. Free Radic Biol Med. 2015 , 4: S0891-5849 . 2. Luu NT, Madden J, Calder PC, Grimble RF, Shearman CP, Chan T, Tull SP, Dastur N, Rainger GE, Nash GB. Comparison of the pro-inflammatory potential of monocytes from healthy adults and those with peripheral arterial disease using an in vitro culture model. Atherosclerosis. 2007 Aug;193(2):259-68. Epub 2006 Sep 18. 3. Change of walking distance in intermittent claudication: impact on inflammation, oxidative stress and mononuclear cells: a pilot study. Dopheide JF, Scheer M, Doppler C, Obst V, Stein P, Vosseler M, Abegunewardene N, Gori T, M\ufcnzel T, Daiber A, Radsak MP, Espinola-Klein C. Clin Res Cardiol. 2015 Mar 15

Lysophosphatidylcholine and Carotid Intima-Media Thickness in Young Smokers: A Role for Oxidized LDL-Induced Expression of PBMC Lipoprotein-Associated Phospholipase A2?

BACKGROUND: Although cigarette smoking has been associated with carotid intima-media thickness (CIMT) the mechanisms are yet not completely known. Lysophosphatidylcholine (lysoPC), a main product of lipoprotein-associated phospholipase A2 (Lp-PLA2) activity, appears to be a major determinant of the pro-atherogenic properties of oxidized LDL (oxLDL) and to induce proteoglycan synthesis, a main player in intimal thickening. In this study we assessed whether cigarette smoking-induced oxidative stress may influence plasma Lp-PLA2 and lysoPC and Lp-PLA2 expression in peripheral blood mononuclear cells (PBMC), as well as the relationship between lysoPC and CIMT. METHODS/RESULTS: 45 healthy smokers and 45 age and sex-matched subjects participated in this study. Smokers, compared to non-smokers, showed increased plasma concentrations of oxLDL, Lp-PLA2 and lysoPC together with up-regulation of Lp-PLA2 (mRNA and protein) expression in PBMC (P<0.001). Plasma Lp-PLA2 positively correlated with both lysoPC (r=0.639, P<0.001) and PBMC mRNA Lp-PLA2 (r=0.484, P<0.001) in all subjects. Moreover CIMT that was higher in smokers (P<0.001), positively correlated with lysoPC (r=0.55, P<0.001). Then in in vitro study we demonstrated that both oxLDL (at concentrations similar to those found in smoker’s serum) and oxidized phospholipids contained in oxLDL, were able to up-regulate mRNA Lp-PLA2 in PBMC. This effect was likely due, at least in part, to the enrichment in oxidized phospholipids found in PBMC after exposure to oxLDL. Our results also showed that in human aortic smooth muscle cells lysoPC, at concentrations similar to those found in smokers, increased the expression of biglycan and versican, two main proteoglycans. CONCLUSIONS: In smokers a further effect of raised oxidative stress is the up-regulation of Lp-PLA2 expression in PBMC with subsequent increase of plasma Lp-PLA2 and lysoPC. Moreover the correlation between lysoPC and CIMT together with the finding that lysoPC up-regulates proteoglycan synthesis suggests that lysoPC may be a link between smoking and intimal thickening