Evaluation of selected parameters of inflammation, coagulation system, and formation of extracellular neutrophil traps (NETs) in the perioperative period in patients undergoing endovascular treatment of thoracoabdominal aneurysm with a branched device (t-Branch).

Extracellular Neutrophils Traps (NETs) and their formation, known as NETosis, have become pivotal in the pathogenesis of aortic aneurysm development. This study investigates the NETosis markers with the assessment of selected parameters of inflammation and coagulation system in patients with thoracoabdominal aortic aneurysms in the pre-and postop period undergoing t-Branch stent-graft implantation. The study included 20 patients with thoracoabdominal aortic aneurysms. Three markers double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), and citrullinated H3 histones (Cit-H3) were tested at three-time points from patients' blood. The parameters of NETosis, inflammation, and coagulation system were examined in the preoperative period (within 24 h before surgery) and in the postoperative period (on the 3rd and 5th postoperative day). Free-circulating DNA (cfDNA) was isolated from the blood using the MagMAXTM Cell-Free DNA Extraction Kit. Double-stranded DNA (dsDNA) and single-stranded DNA (ssDNA) were then quantified using the Qubit dsDNA HS Assay Kit and the Qubit ssDNA Assay Kit. Cit-H3 concentration was determined by enzyme immunoassay ELISA (Cayman). The results revealed the significance of NETs secretion in response to the complex processes after stent-graft implantation. All NET markers increased shortly after surgery, with histones being the first to return to preoperative levels. The lack of normalization of dsDNA and ssDNA levels to preoperative levels by the last postoperative blood collection demonstrates NETs reorganization. The increase in the number of neutrophils was not related to the expansion of postoperative NETosis. The study reveals a new marker of NETosis, ssDNA, that has not been studied so far. The implantation of a stent graft in a patient with TAAA triggers an inflammatory response manifested by an increase in inflammatory parameters. One of the hallmarks of inflammation is the activation of neutrophil extracellular traps.

Development of a novel method for the simultaneous detection of trimethylamine N-oxide and creatinine in the saliva of patients with chronic kidney disease - Its utility in saliva as an alternative to blood.

Chronic kidney disease (CKD) is associated with increased levels of creatinine and other uremic toxins (UTs), which impaired kidneys cannot filtrate. Typically, CKD is diagnosed by calculating the estimated glomerular filtration rate using serum creatinine or cystatin C levels. In pursuit of more sensitive and reliable biomarkers of kidney dysfunction, scientific attention has turned towards other UTs, such as trimethylamine N-oxide (TMAO), successfully quantified in standard matrices, blood and urine. However, less invasive monitoring of kidney function can be performed using an alternative diagnostic biofluid, saliva, which has been shown to contain clinically relevant concentrations of renal function markers. Accurate quantitative estimation of serum biomarkers using saliva measurements can only be achieved provided that there is a tight saliva-serum correlation for the analyte of interest. Therefore, we aimed to verify the correlation between saliva and serum levels of TMAO in CKD patients using newly developed and validated quantitative liquid chromatography coupled to mass spectrometry (LC-MS) method for simultaneous detection of TMAO, and creatinine - the conventional marker of renal impairment. Secondly, we applied this method to quantify TMAO and creatinine levels in the resting saliva of CKD patients collected with a standardised method involving swab-based collectors. A good linear correlation was obtained between the concentration of creatinine in serum and resting saliva of CKD patients (r = 0.72, p = 0.029) and even better in the case of TMAO (r = 0.81, p = 0.008). The analysed validation criteria were fulfilled. No significant influence of the type of swab in the Salivette® device on creatinine and TMAO concentrations in saliva was detected. Our study indicates that saliva can be successfully used in the non-invasive monitoring of renal failure in CKD by measuring salivary TMAO concentrations.

Evaluation of Long-Term Outcomes of Direct Acting Antiviral Agents in Chronic Kidney Disease Subjects: A Single Center Cohort Study.

BACKGROUND: The chronic kidney disease (CKD) population, including kidney transplant recipients (KTRs) and subjects on renal replacement therapy, is particularly vulnerable to unfavorable outcomes from chronic hepatitis C (CHC). Currently, there are oral direct-acting antiviral agents (DAAs) available to eradicate the virus with favorable short-term outcomes; however, their long-term effects are lacking. The aim of the study is to assess the long-term efficacy and safety of DAA therapy in the CKD population. METHODS: An observational, cohort single-center study was performed. Fifty-nine CHC subjects with CKD, treated with DAAs between 2016 and 2018, were enrolled in the study. Safety and efficacy profiles were assessed, including sustained virologic response (SVR), occult hepatitis C infection (OCI) incidence, and liver fibrosis. RESULTS: SVR was achieved in 96% of cases (n = 57). OCI was diagnosed only in one subject following SVR. Significant liver stiffness regression was observed 4 years after SVR compared to baseline values (Mdn = 6.1 kPa, IQR = 3.75 kPa; 4.9 kPa, IQR = 2.9 kPa), p < 0.001. The most common adverse events were anemia, weakness, and urinary tract infection. CONCLUSION: DAAs provide a safe and effective cure for CHC in both CKD patients and KTRs with a favorable safety profile in the long-term follow-up.

NR3C1 Glucocorticoid Receptor Gene Polymorphisms Are Associated with Membranous and IgA Nephropathies.

Glomerular diseases (GNs) are responsible for approximately 20% of chronic kidney diseases. Glucocorticoid receptor gene (NR3C1) single nucleotide polymorphisms (SNPs) are implicated in differences in predisposition to autoimmunity and steroid sensitivity. The aim of this study was to evaluate the frequency of the NR3C1 SNPs-rs6198, rs41423247 and rs17209237-in 72 IgA nephropathy (IgAN) and 38 membranous nephropathy (MN) patients compared to 175 healthy controls and to correlate the effectiveness of treatment in IgAN and MN groups defined as a reduction of proteinuria <1 g/24 h after 12 months of treatment. Real-time polymerase chain reactions and SNP array-based typing were used. We found significant rs41423247 association with MN (p = 0.026); a significant association of rs17209237 with eGFR reduction after follow-up period in all patients with GNs (p = 0.021) and with the degree of proteinuria after 1 year of therapy in all patients with a glomerulopathy (p = 0.013) and IgAN (p = 0.021); and in the same groups treated with steroids (p = 0.021; p = 0.012). We also observed the association between rs41423247 and IgAN histopathologic findings (p = 0.012). In conclusion, our results indicate that NR3C1 polymorphisms may influence treatment susceptibility and clinical outcome in IgAN and MN.

Evaluation of Salivary Indoxyl Sulfate with Proteinuria for Predicting Graft Deterioration in Kidney Transplant Recipients.

Acute kidney injury (AKI) is a significant risk factor for developing chronic kidney disease and progression to end-stage renal disease in elderly patients. AKI is also a relatively common complication after kidney transplantation (KTx) associated with graft failure. Since the lifespan of a transplanted kidney is limited, the risk of the loss/deterioration of graft function (DoGF) should be estimated to apply the preventive treatment. The collection of saliva and urine is more convenient than collecting blood and can be performed at home. The study aimed to verify whether non-invasive biomarkers, determined in saliva and urine, may be useful in the prediction of DoGF in kidney transplant recipients (KTRs) (n = 92). Salivary and serum toxins (p-cresol sulfate, pCS; indoxyl sulfate, IS) concentrations were determined using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Urinary proteins, hemoglobin, and glucose were measured using a semi-quantitative strip test. Salivary IS (odds ratio (OR) = 1.19), and proteinuria (OR = 3.69) were demonstrated as independent factors for the prediction of DoGF. Satisfactory discriminatory power (area under the receiver operating characteristic curve (AUC) = 0.71 ± 0.07) and calibration of the model were obtained. The model showed that categories of the increasing probability of the risk of DoGF are associated with the decreased risk of graft survival. The non-invasive diagnostic biomarkers are a useful screening tool to identify high-risk patients for DoGF.

Selected Clinical Features Fail to Predict Inflammatory Gene Expressions for TNF-α, TNFR1, NSMAF, Casp3 and IL-8 in Tendons of Patients with Rotator Cuff Tendinopathy.

The pathophysiology of rotator cuff tendinopathy is not fully understood, particularly in terms of the local inflammatory process. This study aimed to investigate the expression of selected molecules in the tumour necrosis factor (TNF)-α transduction pathway, including TNF-α, TNF receptor 1 (TNFR1), neutral sphingomyelinase activation associated factor (NSMAF), caspase 3 (Casp3), and interleukin (IL)-8, in patients with rotator cuff tendinopathy that had undergone surgical treatment. We included 44 participants that underwent arthroscopy, due to rotator cuff tendinopathy. Samples from the injured tendon were collected during arthroscopy, and RT-PCR was performed to determine gene expression. Pearson correlation analyses or U-Mann-Whitney test were performed to identify associations with the following parameters: sex, age at admission, body mass index, the presence of night pain, previous treatment (nonsteroidal anti-inflammatory drugs and/or steroids), medical history of the shoulder injury, upper subluxation of the humeral head, and the number of tendons injured. RT-PCR showed that the selected pro-inflammatory factors involved in the TNF-α signalling pathway expression levels were expressed in the tendon tissues. However, the levels of expression varied from patient to patient. Variations were over 250-fold for TNF-α, about 130-fold for TNFR1, NSMAF, and Casp3, and 1000-fold for IL-8. We could not confirm that any of the clinical parameters investigated were associated with the level of gene expression in the TNF-α pathway and IL-8.

Depression But Not Inflammatory Markers Are Increased in Family Versus Nonfamily Caregivers of Persons With Dementia.

We investigated quality of life (WHOQoL-BREF), perceived stress (PSS-10), anxiety and depression (HADS-M), life satisfaction (SWLS), and serum levels of interleukin-6 (IL-6), C-reactive protein (CRP), and cortisol in family caregivers (n = 94) and professional caregivers (n = 48) of demented patients, as well as among noncaregivers (n = 30). Compared with professional caregivers, family caregivers had higher scores in HADS-M depression (P = .003) and anxiety (P = .033), lower life satisfaction (P = .04), and lower quality of life in psychological (P = .02) and social relationship (P = .03) domains. There were no differences in serum levels of IL-6, CRP, or cortisol between caregivers and control participants. In multivariable analysis, when family relationship was considered together with the time period of caregiving and results of the Mini-Mental State Examination test in care recipients (n = 118, 12.49 ± 7.99), only family relationship influenced scores in HADS-M depression (P = .004), SWLS scores (P = .011), and WHOQoL-BREF scores in psychological (P = .011) and social relationship (P = .008) domains. In conclusion, family caregivers are more stressed and have deeper depressive and anxiety disorders, lower life satisfaction, and lower quality of life than professional caregivers.

Epstein-Barr Virus and Human Adenovirus Viremia in Renal Tumors Is Associated with Histological Features of Malignancy.

BACKGROUND: There is growing evidence that viral infections may impact the risk and clinical course of malignancies, including solid tumors. The aim of this study was to assess the possible association of selected chronic/latent viral infections with the clinical course of renal cell carcinoma (RCC). METHODS: In this prospective study we enrolled 27 patients undergoing partial or radical nephrectomy due to the histologically confirmed RCC and followed them up for one year post-operation. Isolation of the nucleic acids was performed using the NucleoSpin Tissue Kit (Macherey-Nagel, Düren, Germany) from tumor tissue and using the EZ1 Virus Mini Kit v2.0 from plasma. The number of viral copies of human adenovirus (ADV), herpes simplex virus HSV-1 and HSV-2, Epstein-Barr virus (EBV), cytomegalovirus (CMV), BK virus (BKV) and John Cunningham virus (JCV) in the tissue and plasma was assessed with real-time PCR. RESULTS: Viral infections were diagnosed in ten patients (37.0%), including three ADV cases (11.1%) and eight EBV cases (29.6%). Infected patients tended to be significantly older (71.3 vs. 57.6 years, p < 0.05), more commonly presented with chronic renal disease (OR 2.4, p < 0.05), diabetes (OR 4.2, p < 0.05) and overweight (OR 2.0, p < 0.05). Regarding oncological data, infected patients were found to have a higher rate of high-grade cancers (OR 5.0, p < 0.05) and a higher rate of papillary RCCs (OR 8.3, p < 0.05). Status of viral infections had no influence on the clinical cancer stage, surgical procedure or survival. CONCLUSIONS: EBV and ADV infections are common in renal cancer patients and increase the risk of high-grade RCC presence. While there is no significant impact on short term survival, further studies are needed to assess the relevance of these findings in a long run.

Prevalence and molecular epidemiology of CMV and EBV among nurses working in pediatrics, transplantology, and primary health care.

OBJECTIVE: The aim of the study was to determine and compare the prevalence of cytomegalovirus (CMV) and Epstein-Barr virus (EBV) antibodies and DNA among nurses working in different profiles of healthcare activity. MATERIAL AND METHODS: The study population comprised 120 women (90 exposed healthcare workers and 30 controls). Blood samples were investigated using chemiluminescent microparticle immunoassays (CMIA) tests to detect the presence of EBV VCA IgM, IgG, and CMV IgM, IgG. Plasma CMV and EBV DNA levels were assessed using real-time polymerase chain reaction (PCR). RESULTS: CMV IgG antibodies were present in 87.80% nurses (86.70% in controls), EBV IgG were present in all the nurses studied and in the control group. No statistically significant differences were noted between the subgroups of nurses and the control group as regards IgG CMV, VCA IgG EBV. CMV IgM/EBV IgM antibodies were negative in all the nurses. CMV/EBV DNA was reported only in the study group. It was not found in any of control group participants. CONCLUSIONS: The positive PCR CMV/EBV markers only in the study group can be indicative of the exposure of nurses to these pathogens being greater than in other people not being professionally involved in patient care. In addition, it was observed that the level of CMV IgG antibodies as well as EBV VCA IgG antibodies tended to be linked to the age and the length of work of nurses working in pediatrics.

Development of the LC-MS/MS method for determining the p-cresol level in plasma.

p-Cresol is a protein-bound uremic retention solute that originates in the intestine through bacterial metabolism and accumulates throughout the body in case of kidney failure. To date, there has been no method to analyze unconjugated p-cresol concentration in the blood with a limit of detection lower than 75 pg. Thus, the aim of this study was to develop and validate a novel liquid chromatography-tandem mass spectrometry method for the determination of unconjugated p-cresol in plasma with a lower detection limit than what has been determined using previously described methods. Sample preparation included derivatization of p-cresol with dansyl chloride (derivatization reagent) showed to be a better approach to analyze the compound. The method optimization involved various pH, time of the reaction, and concentration of derivatization reagent. The validation process was performed according to the procedures prescribed by the European Medicines Agency. All analyzed validation criteria were fulfilled. The novel validated method was applied to compare the level of p-cresol in patients with chronic renal failure before and after dialysis (n = 24). Additionally, the concentration of p-cresol was determined in patients with multiple organ dysfunction syndrome (n = 23). The established method can be used for determination of p-cresol in the plasma in further clinical research.

Protein-Bound Solute Clearance During Hemodialysis.

Indoxyl sulfate (IS) and p-cresol sulfate (p-CS) are protein-bound solutes that accumulate in the blood serum in chronic kidney disease and have a detrimental effect on the kidney and other organs' function. This study seeks to define the effectiveness of IS and p-CS clearance after single dialysis sessions and after 8-week-long cycles of hemodialysis using the following different dialysis modalities in succession: low-flux hemodialysis (lfHD), high-flux hemodialysis (hfHD), and post-dilution hemodiafiltration (HDF). We also investigated to what extent IS and p-CS serum content would associate with some other biochemical indices in patients with chronic kidney diseases. The study included 21 uremic patients. We found that a single session of each modality effectively decreased the content of both IS and p-CS, with the predominance of p-CS decrease. There were no appreciable differences depending on the modality of hemodialysis chosen. However, the leaching effect tended to wear off with the weeks' long dialysis cycles. We further found that a greater inflammation-prone level of hsCRP evoked by dialysis led to a greater removal of solutes, and thus their decrease in the serum, during a single dialysis session. Reversely, a greater protein level might result in a greater solute binding and a decrease in removal. We conclude that there are no major differences in the serum clearance of IS and p-CS depending on the dialysis modality. These protein-bound toxins are significantly cleared from the serum already during the first dialysis session, but their level tends to revert during weeks' long dialysis sessions.

The influence of heavy physical effort on proteolytic adaptations in skeletal and heart muscle and aorta in rats.

Physical effort can elicit differential adaptive changes in the tissues of trained versus untrained rats. Proteolytic activity in the extracellular matrix could be engaged in such adaptation due to its influence on the elasticity of tissues. The effects were investigated of a single physical effort on the activity of elastase, cathepsin K, and plasmin in the skeletal muscles, heart muscles, and aortas of untrained (UT, n=30) and trained (T, n=30) rats. T rats underwent 6 weeks of endurance training. After the last training session, T and UT rats were divided randomly into 3 subgroups. Ten rats from each group (Tpre, n=10) and (UTpre, n=10) were sacrificed. The other 20 rats from each group performed 60 min. of aerobic exercise and were sacrificed immediately post exercise (T0h, n=10; UT0h, n=10) or 3h later (T3h, n=10; UT3h, n=10). Enzyme activity was measured fluorometrically. Cathepsin K and plasmin activity increased in the soleus muscles of UT0h versus UTpre, plasmin activity increased also in UT3h versus UTpre. Elastase, cathepsin K and plasmin activity increased in the heart muscles of T0h and T3h versus Tpre. No aortic differences were observed. Thus, a single bout of physical effort elicited different responses in tissues of T versus UT rats. Increased proteolytic enzyme levels in muscles could influence tissue remodeling. Unchanged aortic cathepsin K levels may help prevent aortic remodeling and neointima formation.

Human cytomegalovirus and Epstein-Barr virus infections increase the risk of death in patients with head and neck cancers receiving radiotherapy or radiochemotherapy.

Viral infections, including cytomegalovirus (CMV) and Epstein-Barr virus (EBV), play an important role in carcinogenesis and can influence patients' prognosis and condition during cancer treatment.The goal of this study was to investigate CMV and EBV infections in patients receiving radiotherapy or radiochemotherapy due to head and neck cancers to determine the influence of these infections on the risk of death. The observation period was 2 years.Of 41 patients enrolled, 11 received radiotherapy (simultaneous-integrated boost intensity-modulated radiation therapy [SIB-IMRT], 2.25 Gy/fraction, 30 fractions, [n = 7] or IMRT, 2 Gy per fraction, 35 fractions, [n = 4]) and 30 received radiochemotherapy (cisplatin 100 mg/m and SIB-IMRT [n = 13] or IMRT [n = 17]). Plasma CMV and EBV DNA levels were assessed using real-time PCR before or during treatment or 4 weeks posttreatment.The risk of death in the group positive for plasma CMV or EBV deoxyribonucleic acid (DNA) was significantly higher compared to the group without detectable plasma CMV (odd ratio [OR]: 7.5, 95% confidence interval [CI]: 1.11-50.67) or EBV DNA (OR: 10.91, 95% CI: 1.135-104.8). Results were confirmed using the Bayesian method. Plasma positivity for CMV or EBV DNA was associated with a higher risk of death (both P = .04).Viral infections negatively affect the survival of patients with head and neck cancers. Diagnosing and treating these viral infections in patients with positive results should be considered.

Chronic, low-dose TMAO treatment reduces diastolic dysfunction and heart fibrosis in hypertensive rats.

Several studies have suggested negative effects of trimethylamine oxide (TMAO) on the circulatory system. However, a number of studies have shown protective functions of TMAO, a piezolyte and osmolyte, in animals exposed to high hydrostatic and/or osmotic stress. We evaluated the effects of TMAO treatment on the development of hypertension and its complications in male spontaneously hypertensive rats (SHRs) maintained on water (SHR-Water) and SHRs drinking TMAO water solution from weaning (SHR-TMAO). Wistar-Kyoto (WKY) rats were used as normotensive controls to discriminate between age-dependent and hypertension-dependent changes. Telemetry measurements of blood pressure were performed in rats between the 7th and 16th weeks of life. Anesthetized rats underwent echocardiographic, electrocardiographic, and direct left ventricular end-diastolic pressure (LVEDP) measurements. Hematoxylin and eosin as well as van Gieson staining for histopathological evaluation were performed. Plasma TMAO measured by chromatography coupled with mass spectrometry was significantly higher in the SHR-Water group compared with the WKY group (~20%). TMAO treatment increased plasma TMAO by four- to fivefold and did not affect the development of hypertension in SHRs. Sixteen-week-old rats in the SHR-Water and SHR-TMAO groups (12-wk TMAO treatment) showed similar blood pressures, angiopathy, and cardiac hypertrophy. However, the SHR-TMAO group had lower plasma NH2-terminal pro-B-type natriuretic peptide, LVEDP, and cardiac fibrosis. In contrast to age-matched WKY rats, 60-wk-old SHRs showed hypertensive angiopathy and heart failure with preserved ejection fraction. Compared with the SHR-Water group, the SHR-TMAO group (56-wk TMAO treatment) showed significantly lower plasma NH2-terminal pro-B-type natriuretic peptide and vasopressin, significantly lower LVEDP, and cardiac fibrosis. In conclusion, a four- to fivefold increase in plasma TMAO does not exert negative effects on the circulatory system. In contrast, increased dietary TMAO seems to reduce diastolic dysfunction in pressure-overloaded hearts in rats. NEW & NOTEWORTHY Chronic, low-dose trimethylamine oxide (TMAO) treatment that increases plasma TMAO by four- to fivefold reduces plasma NH2-terminal pro-B-type natriuretic peptide and vasopressin, left ventricular end-diastolic pressure, and cardiac fibrosis in pressure-overloaded hearts in hypertensive rats. Our study provides evidence that a moderate increase in plasma TMAO does not have a negative effect on the circulatory system. In contrast, increased dietary TMAO seems to reduce diastolic dysfunction in the pressure-overloaded heart.

The utility of saliva testing in the estimation of uremic toxin levels in serum.

Background p-Cresol sulfate (pCS) and indoxyl sulfate (IS) are uremic toxins, high concentrations of which are related to renal failure progression. Saliva could become the first-line diagnostic sample of choice, especially for monitoring purposes. Recently, a method for determination of pCS and IS in saliva was developed. Since no data exist on correlations between the levels of toxins in saliva and serum, the applicability of saliva as a diagnostic material is yet to be established. Here, we present a study on the assessment of the utility of saliva testing in the estimation of uremic toxin levels in serum. Methods The study material included serum and unstimulated, fasting saliva obtained from healthy volunteers (n=26) and patients at all stages of chronic kidney diseases (CKD, n=93). The concentration of pCS and IS in saliva and serum (total and unbound fractions) was determined. The daytime variation of the toxins was studied. Results A correlation was found between pCS and IS in saliva and biological active fractions in serum (0.74; 0.81). The variation of the serum/saliva ratio during the day was negligible, with a median of 10% for pCS and 6% for IS, making saliva a reliable material for the estimation of the uremic toxins in circulation at any time of the day. Significant correlations were observed between salivary toxin levels and estimated glomerular filtration rate (pCS: -0.61; IS: -0.70) as well as significant differences in toxin levels between the stages of CKD. Conclusions Saliva could be a valuable diagnostic material for the estimation of toxin levels in circulation.

Laboratory blood test results beyond normal ranges could not be attributed to healthy aging.

Aging is related to a decline in the function of many organs. The results of blood tests are essential for clinical management and could change over a lifespan reflecting aging. The aim of this study was to examine serum levels of liver, kidney, and bone marrow function and to study their dynamics as a function of age and sex.The cross-sectional study conducted in Poland included 180 healthy individuals (20-90 years) divided into subgroups by sex and decade. These included subgroups of ≥65 or <65 years (men and women). We investigated serum levels of creatinine, estimated glomerular filtration rate, estimated effective renal blood/plasma flow, urine pH, urine neutrophil gelatinase-associated lipocalin (NGAL) as well as serum levels of transaminases, bilirubin, total cholesterol (TC), international normalized ratio (INR), and blood morphology.All parameters were within normal range in all groups. Urine NGAL was higher in men aged ≥65 years than women (25.67 ±â€Š53.65 vs 16.49 ±â€Š34.66, P = .001); serum levels of TC and platelet (PLT) count were higher in women than men aged ≥65 years (221.0 ±â€Š41.7 vs 188.4 ±â€Š48.2 and 250.3 ±â€Š47.8 vs 202.5 ±â€Š57.9, P = .003 and P = .038, respectively). The INR was lower in women (0.97 ±â€Š.06 vs 1.19 ±â€Š0.48, P = .03).These blood tests were normal in healthy people aged ≥65 years. Higher PLT and TC and lower INR in women might indicate a higher risk of cardiovascular diseases. These changes in blood tests were not attributed to aging itself.

Age-dependent increase in serum levels of indoxyl sulphate and p-cresol sulphate is not related to their precursors: Tryptophan and tyrosine.

AIM: Retention of indoxyl sulphate and p-cresol sulphate is associated with many diseases. The aim of the present study was to examine serum levels of indoxyl sulphate and p-cresol sulphate, the dynamics of their changes according to age, and their precursors. METHODS: The study included 180 healthy individuals aged 20-90 years (n = 180), divided into subgroups by decade (n = 30 in each subgroup) and into subgroups of ≥65 years (n = 42) or <65 years (n = 138). Serum indoxyl sulphate and p-cresol sulphate, tryptophan, and tyrosine were measured using high-performance liquid chromatography-mass spectrometry. RESULTS: The 70-90 years age group had higher indoxyl sulphate than the 50-59 years age group (P = 0.033). The 70-90 years age group had higher p-cresol sulphate than the 20-29 years (P < 0.001), 30-39 years (P < 0.001), 40-49 years (P = 0.007) and 50-59 years (P = 0.001) age groups; the 60-69 years age group had higher p-cresol sulphate than the 20-29 years (P = 0.043) and 30-39 years (P = 0.011) age groups. Indoxyl sulphate and p-cresol sulphate serum levels were higher in those aged ≥65 years. Indoxyl sulphate and p-cresol sulphate serum levels correlated positively with age, but not with tryptophan and tyrosine, respectively. CONCLUSIONS: Healthy aging is associated with indoxyl sulphate and p-cresol sulphate serum level increases, which are not linked to tryptophan and tyrosine serum levels. Geriatr Gerontol Int 2017; 17: 1022-1026.

Repeated restraint stress produces acute and chronic changes in hemodynamic parameters in rats.

Noninvasive hemodynamic measurements in rats require placing animals in restrainers. To minimize restraint stress-induced artifacts several habituation protocols have been proposed, however, the results are inconclusive. Here, we evaluated if a four-week habituation is superior to a shorter habituation, or no habituation. This is the first study comparing different habituation protocols with the use of four-week continuous telemetry measurements. We did the experiments on male, 16-week old, Sprague-Dawley rats. Continuous recordings of mean arterial blood pressure (MABP) and heart rate (HR) were made before and during habituation protocols. Rats were subjected either to control (four weeks of restraint-free recordings, n = 5) or two-week (seven restraints, n = 6) or four-week (14 restraints, n = 6) restraint sessions. The restraint protocols included placement of rats in the middle of the dark phase into plastic restrainers as used for tail-cuff measurements. Restraint lasted for 60 min, and was repeated every second day. Each restraint significantly increased MABP (by 15-25 mmHg) and HR (by 40-120 beats/min). Exposure to the restraint protocols decreased diurnal variation in MABP. There was no hemodynamic adaptation to repeated restraint, and no significant difference in hemodynamic response to restraint among controls, the two-week and the four-week groups. In conclusion, our study indicates that measurements in restrained rats are not likely being made without stress-induced changes in MABP. Moreover, in hemodynamic studies in repeatedly restrained rats longer habituation is not superior to shorter habituation.

Training differentially regulates elastin level and proteolysis in skeletal and heart muscles and aorta in healthy rats.

Exercise induces changes in muscle fibers and the extracellular matrix that may depend on elastin content and the activity of proteolytic enzymes. We investigated the influence of endurance training on the gene expression and protein content and/or activity of elastin, elastase, cathepsin K, and plasmin in skeletal and heart muscles and in the aorta. Healthy rats were randomly divided into untrained (n=10) and trained (n=10; 6 weeks of endurance training with increasing load) groups. Gene expression was evaluated via qRT-PCR. Elastin content was measured via enzyme-linked immunosorbent assay and enzyme activity was measured fluorometrically. Elastin content was significantly higher in skeletal (P=0.0014) and heart muscle (P=0.000022) from trained rats versus untrained rats, but not in the aorta. Although mRNA levels in skeletal muscle did not differ between groups, the activities of elastase (P=0.0434), cathepsin K (P=0.0343) and plasmin (P=0.000046) were higher in trained rats. The levels of cathepsin K (P=0.0288) and plasminogen (P=0.0005) mRNA were higher in heart muscle from trained rats, but enzyme activity was not. Enzyme activity in the aorta did not differ between groups. Increased elastin content in muscles may result in better adaption to exercise, as may remodeling of the extracellular matrix in skeletal muscle.

Activity of Proteolytic Enzymes and Level of Cystatin C in the Peripartum Period.

Objectives. The aim of the study was to evaluate the activity of cathepsin B, collagenases, trypsin, and plasmin and concentration of cystatin C in serum of healthy pregnant women in peripartum period. Study Design. The study group included 45 women in uncomplicated pregnancies. Blood samples were collected in four time points. Enzyme activity was measured by spectrofluorometric method. The level of cystatin C was measured using immunonephelometric method. Results. Mean activity of cathepsin B and the level of serum cystatin C were significantly higher in the study group. Collagenase activity was significantly lower in the study group than the control group. No differences in collagenase, plasmin, and trypsin activity on each day of the peripartum period were found. Conclusion. High activity of cathepsin B and increased level of cystatin C are typical for women in late pregnancy. Those levels significantly decrease after delivery which can be associated with potential role of those markers in placental separation. The insignificant changes of cystatin C level in the peripartum period seem to exclude the possibility of using cystatin C as a marker for renal insufficiency in the peripartum period but additional research is necessary to investigate the matter further.