Serum ACE2 Level is Associated With Severe SARS-CoV-2 Infection: A Cross-Sectional Observational Study.

Objectives: Angiotensin-converting enzyme 2 (ACE2) represents the primary receptor for SARS-CoV-2 to enter endothelial cells, causing coronavirus disease of 2019 (COVID-19). In this study, we investigate the association between circulating ACE2 levels with the severity of COVID-19. Methods: Serum ACE2 levels were measured in 144 COVID-19-positive subjects at hospital admission, and 123 COVID-19-negative control subjects. The association between ACE2 and clinical outcomes was analyzed. Results: About 144 COVID-19 patients and 123 healthy controls data were analyzed, the mean age of patients was 62 years and 50% of them were males. The mean age of the control group was 55 years and 63% were males. ACE-II level was measured and compared between COVID-19 patients and controls and revealed no significant differences (P > .05). ACE-II level was measured in COVID-19 patients and compared between different patient's subgroups, ACE II level was not dependent on gender, smoking, ACE intake, or comorbidities (P > .05), and was significantly correlated with cardiovascular diseases (CVS) (P-value = .046) ICU admission (P-value = .0007) and Death (P-value = .0082). Conclusion: There was no significant difference between the COVID-19 and Control group, however, ACE2 serum level was significantly higher in patients with COVID-19 who were critically ill or non-survivors, its increased level is also associated with length of stay. Elevated ACE2 level is associated with the severity of COVID-19 disease, and it has the potential to be a predictor of the severity of the disease.

Evaluating Awareness and Practices Towards Diabetes and Diabetic Retinopathy in Adult Patients Attending the Eye Clinic in a Tertiary Academic Hospital in Jordan.

AIM AND BACKGROUND: Awareness of diabetes mellitus (DM) and its complications, particularly diabetic retinopathy (DR), is one of the main factors of early detection and improved management. This study aims to assess the level of awareness of DM type 2 complications in a cohort of Jordanian patients, with special emphasis on DR. METHODS: A total of 176 consecutive patients with DM type 2 attending the ophthalmology clinic at Jordan University Hospital were included in the study. Each participant responded to a questionnaire which assessed their awareness and behaviors towards DM type 2 and DR. RESULTS: A total of 176 individuals with diabetes responded to the invitation to participate. They were 93 (52.8%) males and 83 (47.2%) females. Mean age (±SEM) for the study population was 60.6 (±0.82) years. Of all participants, 93.8% were aware that diabetes can affect the eyes. Only 4.5% reported that DR could occur without symptoms and/or loss of vision. Symptoms affecting the eyes were the main cause behind attending the ophthalmology clinic in 60.8% of the cases. The awareness score of participants for DM and DR ranged from 4 to 15 out of 20 with a mean score of 11.4/20. Statistically significant relationships of awareness mean score were observed with gender, educational level, employment status, insurance status, Hemoglobin (Hb) A1c level, and dyslipidemia as a co-morbidity (p<0.05). Binary logistic regression revealed disease duration and HbA1c as the main predictive factors of having DR. CONCLUSION: Among this cohort of Jordanian individuals with diabetes, awareness towards DM and DR was relatively low, and patient practices did not correlate with perceived awareness. Awareness scores correlated with HbA1c readings and higher educational levels among other variables. Emphasis on communication strategies and patient education is essential in establishing efficient screening programs and effective strategies to curtail visual impairment and other complications of the diabetes pandemic.

Relationship between Serum Vascular Endothelial Growth Factor Levels and Stages of Diabetic Retinopathy and Other Biomarkers.

AIM: This study aims to measure serum vascular endothelial growth factor (VEGF) levels in a sample of Jordanian patients and to determine their relationship with the different stages of diabetic retinopathy. It also explores the correlation between VEGF concentrations and different biochemical and demographic findings. MATERIALS AND METHODS: A total of 167 adults participated in the study. Participants were divided into two main categories: patients with diabetes mellitus (DM) type 2 without diabetic retinopathy (DR) (N = 62) and patients with DM type 2 affected by DR (N = 105). DR patients were further subclassified into nonproliferative (N = 41) and proliferative (N = 64). Basic laboratory tests were measured to correlate with VEGF levels. Irisin, a hormone linked to diabetic retinopathy was also measured and correlated with VEGF. RESULTS: Serum VEGF was found to positively correlate with the severity of diabetic retinopathy. The means of VEGF serum concentrations were 60 pg/mL for controls, 133 pg/mL for nonproliferative DR patients, and 229 pg/mL for proliferative DR patients. We found a significant positive correlation with glycosylated hemoglobin (HbA1c), and a significant negative correlation with high-density lipoprotein (HDL) levels, age, and irisin. CONCLUSION: In this cohort of Jordanian diabetics, serum VEGF concentrations strongly correlated with the presence and stages of diabetic retinopathy, suggesting it as an appropriate indicator for diabetic retinopathy early detection and management in this society. VEGF levels also significantly correlated with HbA1c, HDL, and irisin levels. Further studies are encouraged to explore these relationships in other ethnic groups and with different diabetic complications.

PKM2 and HIF-1α regulation in prostate cancer cell lines.

Prostate cancer (PCA) is one of the most common cancer types in men, with cancer progression being linked to hypoxia and the induction of hypoxia-inducible factor (HIF).We investigated the expression of pyruvate kinase M2 (PKM2), its regulation by HIF isoforms 1α and 2α, and its role in HIF stabilization. We additionally examined cell survival in the prostate cancer cell lines PC3 and LNCaP under severe hypoxic (0.1% O2) and normoxic (20% O2) conditions. qRT-PCR showed higher up-regulation of PKM2 mRNA expression in LNCaP cells than in PC3 cells, while western blotting showed that PKM2 protein levels were up-regulated only in LNCaP cells. Inhibition of HIF-1α and HIF-2α by small interfering RNA (si-RNA) demonstrated HIF-1α dependent up-regulation of PKM2 at the mRNA and protein levels in LNCaP cells. PKM2 inhibition by si-RNA significantly decreased hypoxia-response element (HRE) activation in a gene reporter assay and down-regulated HIF-1α target vascular endothelial growth factor (VEGF) mRNA expression in PC3 cells, whereas HIF-1α protein levels were not significantly reduced. Additionally, PKM2 inhibition significantly reduced clonogenic survival in both cell lines in a colony formation assay. Prolyl hydroxylase 3 (PHD3) mRNA expression was up-regulated in both cell lines. It has been shown that PKM2 expression is regulated by HIF-1α and that PKM2 favors HIF-1α transactivation under mild (1% O2) but not severe (0.1% O2) hypoxic conditions, and some of our findings are consistent with these previous results. However, this mechanism was not fully observed in our studied cell lines, as PKM2 regulation and HIF-1α stabilization at the transactivation level occurred under severe hypoxic conditions. This discrepancy suggests that tumor tissue origin and cell type influence this model. Our findings expand the current knowledge of the mechanisms of PCA regulation, and would be important in developing novel therapeutic strategies.

Carboxyl group of Glu113 is required for stabilization of the diferrous and bis-Fe(IV) states of MauG.

The diheme enzyme MauG catalyzes a six-electron oxidation required for post-translational modification of a precursor of methylamine dehydrogenase (preMADH) to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. Crystallographic studies have implicated Glu113 in the formation of the bis-Fe(IV) state of MauG, in which one heme is Fe(IV)═O and the other is Fe(IV) with His-Tyr axial ligation. An E113Q mutation had no effect on the structure of MauG but significantly altered its redox properties. E113Q MauG could not be converted to the diferrous state by reduction with dithionite but was only reduced to a mixed valence Fe(II)/Fe(III) state, which is never observed in wild-type (WT) MauG. Addition of H2O2 to E113Q MauG generated a high valence state that formed more slowly and was less stable than the bis-Fe(IV) state of WT MauG. E113Q MauG exhibited no detectable TTQ biosynthesis activity in a steady-state assay with preMADH as the substrate. It did catalyze the steady-state oxidation of quinol MADH to the quinone, but 1000-fold less efficiently than WT MauG. Addition of H2O2 to a crystal of the E113Q MauG-preMADH complex resulted in partial synthesis of TTQ. Extended exposure of these crystals to H2O2 resulted in hydroxylation of Pro107 in the distal pocket of the high-spin heme. It is concluded that the loss of the carboxylic group of Glu113 disrupts the redox cooperativity between hemes that allows rapid formation of the diferrous state and alters the distribution of high-valence species that participate in charge-resonance stabilization of the bis-Fe(IV) redox state.

A Trp199Glu MauG variant reveals a role for Trp199 interactions with pre-methylamine dehydrogenase during tryptophan tryptophylquinone biosynthesis.

MauG catalyzes posttranslational modifications of a methylamine dehydrogenase precursor (preMADH) to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. Trp199 is present at the site of interaction between MauG and preMADH and is critical to this process as it mediates hole hopping during the inter-protein electron transfer that is required for catalysis. Trp199 was converted to Glu and the structure and reactivity of the W199E/preMADH complex were characterized. The results reveal that the nature of residue 199 is also important for productive complex formation between preMADH and MauG.

Effects of the loss of the axial tyrosine ligand of the low-spin heme of MauG on its physical properties and reactivity.

MauG catalyzes posttranslational modifications of methylamine dehydrogenase to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. MauG possesses a five-coordinate high-spin and a six-coordinate low-spin ferric heme, the latter with His-Tyr ligation. Replacement of this tyrosine with lysine generates a MauG variant with only high-spin ferric heme and altered spectroscopic and redox properties. Y294K MauG cannot stabilize the bis-Fe(IV) redox state required for TTQ biosynthesis but instead forms a compound I-like species on reaction with peroxide. The results clarify the role of Tyr ligation of the five-coordinate heme in determining the physical and redox properties and reactivity of MauG.

Functional importance of tyrosine 294 and the catalytic selectivity for the bis-Fe(IV) state of MauG revealed by replacement of this axial heme ligand with histidine .

The diheme enzyme MauG catalyzes the posttranslational modification of a precursor protein of methylamine dehydrogenase (preMADH) to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. It catalyzes three sequential two-electron oxidation reactions which proceed through a high-valent bis-Fe(IV) redox state. Tyr294, the unusual distal axial ligand of one c-type heme, was mutated to His, and the crystal structure of Y294H MauG in complex with preMADH reveals that this heme now has His-His axial ligation. Y294H MauG is able to interact with preMADH and participate in interprotein electron transfer, but it is unable to catalyze the TTQ biosynthesis reactions that require the bis-Fe(IV) state. This mutation affects not only the redox properties of the six-coordinate heme but also the redox and CO-binding properties of the five-coordinate heme, despite the 21 Å separation of the heme iron centers. This highlights the communication between the hemes which in wild-type MauG behave as a single diheme unit. Spectroscopic data suggest that Y294H MauG can stabilize a high-valent redox state equivalent to Fe(V), but it appears to be an Fe(IV)═O/π radical at the five-coordinate heme rather than the bis-Fe(IV) state. This compound I-like intermediate does not catalyze TTQ biosynthesis, demonstrating that the bis-Fe(IV) state, which is stabilized by Tyr294, is specifically required for this reaction. The TTQ biosynthetic reactions catalyzed by wild-type MauG do not occur via direct contact with the Fe(IV)═O heme but via long-range electron transfer through the six-coordinate heme. Thus, a critical feature of the bis-Fe(IV) species may be that it shortens the electron transfer distance from preMADH to a high-valent heme iron.

Long-range electron transfer reactions between hemes of MauG and different forms of tryptophan tryptophylquinone of methylamine dehydrogenase.

The diheme enzyme MauG catalyzes the post-translational modification of a precursor protein of methylamine dehydrogenase (preMADH) to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. This six-electron oxidation of preMADH requires long-range electron transfer (ET) as the structure of the MauG-preMADH complex reveals that the shortest distance between the modified residues of preMADH and the nearest heme of MauG is 14.0 A [Jensen, L. M. R., Sanishvili, R., Davidson, V. L., and Wilmot, C. M. (2010) Science 327, 1392-1394]. The kinetics of two ET reactions between MADH and MauG have been analyzed. Interprotein ET from quinol MADH to the high-valent bis-Fe(IV) form of MauG exhibits a K(d) of 11.2 microM and a rate constant of 20 s(-1). ET from diferrous MauG to oxidized TTQ of MADH exhibits a K(d) of 10.1 microM and a rate constant of 0.07 s(-1). These similar K(d) values are much greater than that for the MauG-preMADH complex, indicating that the extent of TTQ maturity rather than its redox state influences complex formation. The difference in rate constants is consistent with a larger driving force for the faster reaction. Analysis of the structure of the MauG-preMADH complex in the context of ET theory and these results suggests that direct electron tunneling between the residues that form TTQ and the five-coordinate oxygen-binding heme is not possible, and that ET requires electron hopping via the six-coordinate heme.