2024 Guidelines of the Polish Society of Laboratory Diagnostics and the Polish Lipid Association on laboratory diagnostics of lipid metabolism disorders.

Lipid disorders are the most common (even 70%) and worst monitored cardiovascular risk factor (only 1/4 of patients in Poland and in CEE countries are on the low-density lipoprotein cholesterol (LDL-C) goal). To improve this, clear and simple diagnostic criteria should be introduced for all components of the lipid profile. These are the updated guidelines of the two main scientific societies in Poland in the area - the Polish Society of Laboratory Diagnostics (PSLD) and the Polish Lipid Association (PoLA), which, in comparison to those from 2020, introduce few important changes in recommendations (two main lipid targets, new recommendations on LDL-C measurements, calculations new goals for triglycerides, new recommendations on remnants and small dense LDL) that should help the practitioners to be early with the diagnosis of lipid disorders and in the effective monitoring (after therapy initiation), and in the consequence to avoid the first and recurrent cardiovascular events.

Lipoprotein(a) is associated with DNA damage in patients with heterozygous familial hypercholesterolemia.

Heterozygous familial hypercholesterolemia (HeFH) is a common autosomal-dominant inherited disorder associated with atherosclerotic cardiovascular disease (ASCVD). HeFH subjects have a higher lipoprotein(a), i.e. Lp(a), concentration than the general population. Patients with FH are exposed to elevated levels of LDL from birth and ox-LDL may induce other oxidation pathways. The aim of the study was to determine the levels of markers of oxidative stress and DNA damage in patients with HeFH and describe the effect of Lp(a) on the resulting damage. Higher DNA damage was identified in patients with HeFH compared to the normolipidemic ones, and ASCVD was associated with greater damage. Oxidative stress markers were elevated in HeFH patients; however, only ox-LDL was higher in the ASCVD group and its level correlated with DNA damage. A positive correlation was found between DNA damage and Lp(a) concentration in the HeFH patients. Higher levels of Lp(a) were associated with greater DNA damage, especially in patients with HeFH and ASCVD. In HeFH patients, the optimal Lp(a) cut-off point associated with ASCVD is > 23.45 nmol/L, i.e. much lower than for the general population; however this cut-off point needs validation in a larger group of HeFH patients.

When the same treatment has different response: The role of pharmacogenomics in statin therapy.

Statins, also known as HMG-CoA reductase inhibitors, are one of the most potently prescribed and thoroughly researched medications, predominantly utilized for managing cardiovascular diseases by modulating serum cholesterol levels. Despite the well-documented efficacy of statins in reducing overall mortality via attenuating the risk of cardiovascular diseases, notable interindividual variability in therapeutic responses persists as such variability could compromise the lipid-lowering efficacy of the drug, potentially increasing susceptibility to adverse effects or attenuating therapeutic outcomes.This phenomenon has catalysed a growing interest in the scientific community to explore common genetic polymorphisms within genes that encode for pivotal enzymes within the pharmacokinetic pathways of statins. In our review, we focus to provide insight into potentially clinically relevant polymorphisms associated with statins' pharmacokinetic participants and assess their consequent implications on modulating the therapeutic outcomes of statins among distinct genetic carrier.

The protective effects of empagliflozin on DNA oxidative changes in a model of vascular endothelial and smooth muscle cells damaged by oxidized cholesterol.

BACKGROUND: Diabetes patients often suffer chronic vascular complications resulting from endothelial dysfunction, smooth muscle cell (SMC) proliferation, inflammation and disturbed oxidative balance. Empagliflozin is one of three approved sodium-glucose cotransporter 2 (SGLT2) inhibitors for type 2 diabetes mellitus. THE AIM OF THIS STUDY: was to determine the protective and repairing effect of EMPA in a model of vascular endothelial and SMC damage with 25-hydroxycholesterol (25-OHC). METHODS: Human umbilical vascular endothelial cells (HUVECs) and SMCs were treated with compounds which induce DNA single-strand breaks (SSBs) and subjected to comet assay. Oxidative DNA damage was detected using endonuclease III (Nth) or human 8 oxoguanine DNA glycosylase (hOOG1). Reactive oxygen species (ROS) formation was determined by the fluorescence of a 6-carboxy-2',7'-dichlorodihydrofluoresce probe in diacetate (H2DCFDA). RESULTS: 25-OHC-stimulated SMCs showed greater resistance to ROS generation and DNA damage compared to HUVECs. In both experimental models, EMPA treatment was associated with lower ROS production and DNA damage, including oxidative damage to purines and pyrimidines. This effect was not dose-dependent. EMPA was found to counteract this DNA damage by inhibiting ROS production. CONCLUSIONS: It appears that the EMPA induced indirect repair of DNA by inhibiting ROS production.

Severe COVID-19 classified by simple covid risk index is associated with higher levels of advanced oxidation protein products and 8-hydroxy 2 deoxyguanosine.

SARS-CoV-2 has become one of the most important and challenging medical research topics in recent years. The presence of endothelial dysfunction, immune thrombosis, and oxidative stress contributes to complications and requires more extended hospitalisation of patients. In this article, we focused on analysing the impact of oxidative stress on the severity of COVID-19 infection. The study group consisted of 72 patients with laboratory-confirmed SARS-CoV enrolled. The patients were divided into moderate and severe diseases according to the SCRI (Simple Covid Risk Index, including lymphocyte/D-dimer ratio). Using the ELISA kit, we determined the level of AOPP and 8-OHdG. Patients with severe COVID-19 had higher levels of both AOPP (P < 0.05) and 8-OHdG (P < 0.05) compared to patients with moderate disease. Albumin levels were significantly lower (P < 0.001), although fibrinogen (P < 0.01), D-dimer (P < 0.001), and TF (P < 0.05) levels were higher in severe patients than in moderate course. AOPP/Alb was also higher among severe patients (P < 0.05). Our data suggest a potential role for AOPP and 8-OHdG in predicting the outcome of SARS-CoV-2 patients. Elevated AOPP levels were associated with increased Dimer-D, TF, and vWF activity levels.

Interleukin-33/sST2: Dynamic assessment in patients with acute coronary syndrome.

PURPOSE: Interleukin (IL)-33 and its soluble receptor ST2 (sST2) play a crucial role in the immune response. sST2 has been approved by the Food and Drug Administration as a prognostic biomarker of mortality in chronic heart failure patients, however, the role of IL-33 and sST2 in atherosclerotic cardiovascular disease remains unclear. The aim of this study was to measure serum level of IL-33 and sST2 of patients at the onset of acute coronary syndrome (ACS) and 3 months after primary percutaneous revascularization. PATIENTS AND METHODS: Forty patients were divided into ST segment elevation myocardial infarction (STEMI) group, non-ST segment elevation myocardial infarction (NSTEMI) and unstable angina (UA) group. IL-33 and sST2 level were measured with ELISA. Additionally, IL-33 expression in peripheral blood mononuclear cells (PBMCs), was evaluated. RESULTS: All ACS patients had a significantly lower level of sST2 3 months after ACS as compared to the baseline (p â€‹< â€‹0.039). The STEMI patients had higher serum levels of IL-33 at the moment of ACS as compared to 3 months after the event, with an average decrease of 17.87 â€‹pg/ml (p â€‹< â€‹0.007). Conversely, sST2 serum levels were still high after 3 months following an ACS in STEMI patients. ROC curve demonstrated that increased IL-33 serum level could be STEMI predictor. CONCLUSIONS: The assessment of the baseline and dynamics of changes in IL-33 and sST2 concentrations in patients with ACS may be important for the diagnostic process and may help in understanding of how the immune mechanisms work at the moment of an ACS event.

SGLT2 Inhibitors May Restore Endothelial Barrier Interrupted by 25-Hydroxycholesterol.

SGLT2 (Sodium-glucose Cotransporter-2) inhibitors are newer glucose-lowering drugs with many cardiovascular benefits that are not fully understood yet. Endothelial integrity plays a key role in cardiovascular homeostasis. 25-hydroxycholesterol (25-OHC), which is a proatherogenic stimuli that impairs endothelial barrier functions. VE-cadherin is an endothelial-specific protein crucial in maintaining endothelial integrity. The aim of this study was to assess the influence of SGLT2i on the integrity of endothelial cells interrupted by 25-OHC. We also aimed to evaluate whether this effect is associated with changes in the levels of VE-cadherin. We pre-incubated HUVECs with 10 µg/mL of 25-hydroxycholesterol (25-OHC) for 4 h and then removed it and incubated endothelial cells with 1 µM of empagliflozin, 1 µM canagliflozin, or 1 µM dapagliflozin for 24 h. The control group included HUVECs cultured with the medium or with 25-OHC 10 µg/mL. The integrity of endothelial cells was measured by the RTCA-DP xCELLigence system, and VE-cadherin was assessed in confocal microscopy. Our results show that SGLT2 inhibitors significantly increase endothelial integrity in comparison to medium controls, and they improve endothelial cell integrity interrupted by 25-OHC. This effect is associated with significant improvements in VE-cadherin levels. SGLT2i: empagliflozin, canagliflozin, and dapagliflozin have a beneficial effect on the endothelial cell integrity and VE-cadherin levels reduced by 25-OHC.

The effect of lipid-lowering therapies on the pro-inflammatory and anti-inflammatory properties of vascular endothelial cells.

Atherosclerosis and cardiovascular events can be prevented, or treated, using statin therapy, either alone or in combination with ezetimibe. Chronic inflammation, vascular proliferation, and the development of atherosclerosis are also influenced by 25-hydroxycholesterol (25-OHC). The aim of the study was to compare the direct pleiotropic effects of two commonly-used statins (atorvastatin, rosuvastatin), ezetimibe, and their combinations, on the mRNA expression of pro-inflammatory IL1ß, IL-18 and IL-23 and anti-inflammatory TGFß, IL-35 (EBI3, IL-12 subunits), IL-10 and IL-37, in endothelial cells damaged by 25-OHC. It also analyzed IL-35 expression at the protein level. HUVECs were stimulated with atorvastatin (5 µM), rosuvastatin (10 µM), ezetimibe (1.22 µM), atorvastatin-ezetimibe (5 µM + 1.22 µM) or rosuvastatin-ezetimibe (10 µM + 1.22 µM), with or without pre-incubation with 10 µg/mL 25-OHC. mRNA expression was analyzed by real-time PCR. The protein level of IL-35 was analyzed by ELISA. In the pre-stimulated HUVECs, atorvastatin and rosuvastatin decreased mRNA expression of IL1ß, IL-18, IL-23, TGFß, IL35 and increased mRNA expression of IL-10 and IL-37 compared to 25-OHC. Furthermore, only incubation with rosuvastatin and rosuvastatin-ezetimibe decreased IL-35 mRNA and protein levels. Ezetimibe down-regulated only IL1ß. Treatment with rosuvastatin-ezetimibe and atorvastatin-ezetimibe reversed the effect of 25-OHC in IL1ß, IL-18 and IL-35 mRNA expression. In conclusion, rosuvastatin has the strongest anti-inflammatory effects and is the best at reducing the effect of oxysterols. Both statins exert a greater anti-inflammatory effect than ezetimibe. The anti-inflammatory effect of the combination therapies appears to be based on the effects of the statins alone and not their combination with ezetimibe.

Searching for serum biomarkers linking coronary heart disease and Helicobacter pylori infection using infrared spectroscopy and artificial neural networks.

Helicobacter pylori (Hp) Gram-negative bacteria cause gastritis or gastric ulcers. They may be involved in the development of systemic diseases i.e. coronary heart disease (CHD). Both Hp infection and CHD are related to inflammation accompanied by C-reactive protein (CRP), tumor necrosis factor alfa (TNF-α) and homocysteine. Low density lipoprotein (LDL) and triglicerides are a classic risk factors of CHD. Infrared spectroscopy has been introduced for monitoring chronic infections or endogenous disorders using specific absorption bands for biocomponents typed as diagnostic markers. In this study we selected specific motives of infrared radiation (IR) spectra for the sera from CHD patients infected with Hp. In total 141 sera were used: 90 from patients with CHD, all Hp positive, and 51 from healthy donors, 32 Hp negative and 21 Hp positive. Hp status was evaluated by anti-Hp IgG antibodies and/or 13C urea breath testing. IR spectra were measured using FT-IR/FT-NIR Spectrum 400 spectrometer (PerkinElmer) chemometrically analyzed using artificial neural networks and they showed differences in absorption bands corresponding to triglicerides, CRP, homocysteine, LDL and TNF-α, and selected component groups between CHD patients infected with Hp vs healthy uninfected donors (96.15% accuracy). Triglicerides and CRP were the best biomarkers linking Hp infection with CHD.

BCG and SARS-CoV-2-What Have We Learned?

Despite controversy over the protective effect of the BCG (Bacille Calmette-Guérin) vaccine in preventing pulmonary tuberculosis (TB) in adults, it has been used worldwide since 1921. Although the first reports in the 1930s had noted a remarkable decrease in child mortality after BCG immunization, this could not be explained solely by a decrease in mortality from TB. These observations gave rise to the suggestion of nonspecific beneficial effects of BCG vaccination, beyond the desired protection against M. tuberculosis. The existence of an innate immunity-training mechanism based on epigenetic changes was demonstrated several years ago. The emergence of the pandemic caused by the severe acute respiratory syndrome coronavirus (SARS-CoV-2) in 2019 revived the debate about whether the BCG vaccine can affect the immune response against the virus or other unrelated pathogens. Due to the mortality of the coronavirus disease (COVID-19), it is important to verify each factor that may have a potential protective value against the severe course of COVID-19, complications, and death. This paper reviews the results of numerous retrospective studies and prospective trials which shed light on the potential of a century-old vaccine to mitigate the pandemic impact of the new virus. It should be noted, however, that although there are numerous studies intending to verify the hypothesis that the BCG vaccine may have a beneficial effect on COVID-19, there is no definitive evidence on the efficacy of the BCG vaccine against SARS-CoV-2.

Elevated Lp(a) and course of COVID-19: Is there a relationship?

BACKGROUND: Lipoprotein (a)-Lp(a) has proinflammatory, prothrombotic and proatherogenic properties and may theoretically influence the course of COVID-19. OBJECTIVES: The aim of the study was to explore whether patients hospitalized due to COVID-19 with Lp(a) ≥30mg/dl may develop a worse course of the disease, increased incidence of thromboembolic complications, intubation and ICU hospitalization or death. PATIENTS AND METHODS: A retrospective analysis was performed of 124 patients hospitalized due to COVID-19 in the Department of Internal Diseases and Clinical Pharmacology between 29 November 2020 and 15 April 2021. The only exclusion criterion was age≥80 years. Patients were divided into two groups: 1. COVID-19 patients with Lp(a) <30mg/dl regarded as not elevated n = 80; 2. COVID-19 patients with Lp(a) ≥30 regarded as elevated n = 44. RESULTS: A total of 124 COVID-19 patients were included in the study (66 men and 58 women) with a mean age of 62.8±11 years. COVID-19 patients with elevated Lp(a) level had significantly longer hospitalization time (11 vs. 9.5 days; p = 0.0362), more extensive radiological changes in CT scan (35 vs. 30%; p = 0.0301) and higher oxygen demand on admission (8 vs. 5L/min; p = 0.0428). Elevated Lp(a) was also associated with significantly higher OR for High Flow Nasal Oxygen Therapy (HFNOT) OR = 3.5 95%CI(1.2;8.9), p = 0.0140, Intubation and ICU OR = 4.1 95%CI(1.1;15.2) p = 0.0423, Death OR = 2.8 95%CI(0.9;8.5), p = 0.0409. CONCLUSIONS: Elevated Lp(a) might be one of the factors which contribute to a more severe course of COVID-19; however, further studies including larger groups of patients are needed.

Antibodies towards TVLLPVIFF Amino Acid Sequence of TNF Receptor Induced by Helicobacter pylori in Patients with Coronary Heart Disease.

BACKGROUND: Molecular mimicry between Helicobacter pylori (Hp) and the host components resulting in induction of cross-reacting antibodies has been suggested as accessory mechanism in the development of coronary heart disease (CHD). A potential target for antibodies induced during Hp infection by the components of these bacteria might be amino acid sequence TVLLPVIFF (P1) of tumor necrosis factor receptor (TNFR), which is exposed on vascular endothelium and immunocompetent cells, driving inflammation. AIM: To examine whether anti-P1 IgG are induced during Hp infection in CHD patients. METHODS: Sera from CHD patients infected with Hp (54) vs. sera of uninfected healthy donors (22) were tested by the ELISA for anti-H. pylori antibodies, anti-P1 IgG, and for antibodies towards control sequence IAKEGFEKIS (P2). Sera of Caviae porcellus infected experimentally with Hp (30) or uninfected (10) were included into this study. The same serum samples, which were positive for anti-P1 IgG, were adsorbed with Hp and then subjected to the ELISA. The biological activity of anti-P1 IgG was assessed in complement (C1q) binding assay. RESULTS: Sera of 43 CHD patients seropositive for anti-Hp IgG contained anti-P1 IgG binding C1q. Additionally, 10 serum samples of animals seropositive for anti-Hp IgG contained anti-P1 IgG. Anti-P1 IgG in tested sera were neutralized by their adsorption with Hp. CONCLUSION: In CHD patients infected with Hp, antibodies cross-reacting with TNFR common sequence are produced. Further studies are necessary to define immunogenic Hp determinants and to confirm possible cellular effects of cross-reacting antibodies.

Expression of anti and pro-inflammatory genes in human endothelial cells activated by 25-hydroxycholesterol: A comparison of rivaroxaban and dabigatran.

Atherosclerosis is associated with a haemostatic imbalance characterized by excessive activation of pro-inflammatory and pro-coagulant pathways. Non-vitamin K antagonists oral anticoagulant (NOACs) may reduce the incidence of cardiovascular events, cerebral ischemia, thromboembolic events and atherosclerosis. Chronic inflammation, vascular proliferation and the development of atherosclerosis is also influenced by 25-hydroxycholesterol (25-OHC). The aim of the study was to assess the effect of rivaroxaban and dabigatran on the messenger RNA (mRNA) expression of anti-inflammatory cytokines transforming growth factor ß (TGF-ß), interleukin (IL)-37, IL-35 as well as of pro-inflammatory cytokines IL-18 and IL-23, in endothelial cells damaged by 25-OHC. Human umbilical vascular endothelial cells (HUVECs) were treated with 25-OHC (10 µg/mL), rivaroxaban (100, 500 ng/mL), dabigatran (100, 500 ng/mL), 25-OHC + rivaroxaban, and 25-OHC + dabigatran. The mRNA expression of TGF-ß, IL-37, IL-35 subunits EBI3 and p35, IL-18, and IL-23 was analysed using real-time polymerase chain reaction (PCR). The results showed that 25-OHC decreased TGF-ß and IL-37 mRNA expression and increased EBI3, p35, IL-18, IL-23 mRNA expression in endothelial cell as compared to an untreated control (P < .05). Messenger RNA expression of TGF-ß and IL-37 significantly increased following stimulation with rivaroxaban and dabigatran as compared to an untreated control (P < .01). In HUVECs pre-treated with oxysterol, rivaroxaban and dabigatran increased mRNA expression of TGF-ß, IL-37 and decreased mRNA expression of EBI3, p35, IL-23 and IL-18 as compared to 25-OHC (P < .01). Our finding suggests that both rivaroxaban and dabigatran inhibit the inflammatory activation caused by oxysterol in vitro.

Euthyroid Sick Syndrome as a Prognostic Indicator of COVID-19 Pulmonary Involvement, Associated With Poorer Disease Prognosis and Increased Mortality.

OBJECTIVE: The prevalence of euthyroid sick syndrome (ESS) and its association with the prognosis of COVID-19 and mortality in patients with lung involvement in COVID-19 have not yet been elucidated. METHODS: Clinical and laboratory data of patients with COVID-19 with or without ESS were collected retrospectively and analyzed on admission. All subjects were admitted to the Department of Internal Diseases and Clinical Pharmacology at Bieganski Hospital between December 2020 and April 2021. RESULTS: In total, 310 medical records of patients with COVID-19 were analyzed retrospectively. Among 215 enrolled patients, 82 cases of ESS were diagnosed. The patients with ESS had higher pro-inflammatory factor levels, longer hospitalizations, and a higher risk of requiring high-flow nasal oxygen therapy or intubation than the patients without ESS. The Kaplan-Meier curve indicated that the patients with ESS had a lower probability of survival when computed tomography showed ≤50% parenchymal involvement compared with that in patients without ESS. However, no differences in mortality were noted in those with more than 50% parenchymal involvement. The survival curve showed that ESS was associated with a higher risk of mortality during hospitalization. CONCLUSION: ESS is closely associated with a poor prognosis, including longer hospitalizations, more frequent intubation, transfer to the intensive care unit, and a higher mortality rate in patients with COVID-19. ESS is a potential prognostic predictor of survival, regardless of lung involvement in COVID-19.

Dabigatran: its protective effect against endothelial cell damage by oxysterol.

Recent data showed that dabigatran can reduce not only procoagulatory effects but also block proinflammatory stimuli by inhibiting the expression of cytokines and chemokines and reducing thrombin-induced endothelial permeability. The aim of our study was to assess the effect of dabigatran on the integrity and inflammatory properties of endothelial cells stimulated by 25-hydroxycholesterol (25-OHC, oxysterol). HUVECs (Human Umbilical Vein Endothelial Cells) were stimulated with 25-hydroxycholesterol 10 µg/ml, dabigatran 100 ng/ml or 500 ng/ml and 25-hydroxycholesterol + dabigatran (100 ng/ml, 500 ng/ml). HUVEC integrity and permeability was measured in the RTCA-DP xCELLigence system and by the paracellular flux system. The mRNA expression of ICAM-1, VEGF, IL-33, MCP-1 and TNF-α was analyzed by Real-time PCR. Cell apoptosis and viability was measured by flow cytometry. VEGF protein concentration was assessed in supernatants by ELISA. VE-cadherin expression in endothelial cells was evaluated by confocal microscopy. Pre-stimulation of HUVECs with 25-OHC decreased endothelial cell integrity (p < 0.001) and increased the expression of IL-33, ICAM-1, MCP-1, VEGF, TNF-α mRNA (p < 0.01) compared to unstimulated controls. Following stimulation of HUVECs with dabigatran 100 ng/ml or 500 ng/ml restored HUVEC integrity interrupted by 25-OHC (p < 0.001). In HUVECs pre-stimulated with oxysterol, dabigatran stimulation decreased mRNA expression of the proinflammatory cytokines IL-33 and TNF-α, chemokines MCP-1 ICAM-1 and VEGF (p < 0.01). Dabigatran 500 mg/ml+ 25-OHC increased the endothelial expression of VE-cadherin as compared to 25-OHC (p < 0.01). Our findings suggest that dabigatran stabilizes the endothelial barrier and inhibits the inflammation caused by oxysterol.