Heart failure (CHF) in combination with chronic kidney disease (CKD) or hypertension worsens the patient's prognosis and results in poor survival rate. In collaboration with colleagues from the Institute for Clinical and Experimental Medicine (Prague), we examined if the addition of endothelin type A (ETA) receptor antagonist to the angiotensin-converting enzyme inhibitor (ACEi) will bring additional beneficial effects in experimental model of volume overload HF that was combined either with chronic kidney disease (induced by 5/6 renal mass reduction - 5/6 NX) in normotensive Hannover Sprague Dawley rats or with hypertension in heterozygous TGR. The treatment with ETA receptor antagonist alone or ACEi alone and the combined treatment similarly improved the survival and cardiac function. However, the combined treatment exerted the best renoprotection, causing additional decrease in albuminuria and reducing renal glomerular and tubulointerstitial injury as compared with ACE inhibition alone.
Kala P, et al. Endothelin type A receptor blockade increases renoprotection in congestive heart failure combined with chronic kidney disease: Studies in 5/6 nephrectomized rats with aorto-caval fistula. Biomed Pharmacother. 2023 Feb;158:114157
Kala P, et al. Endothelin type A receptor blockade attenuates aorto-caval fistula-induced heart failure in rats with angiotensin II-dependent hypertension. J Hypertens. 2023 Jan 1;41(1):99-114.
In collaboration with dr. Maletínská group from IOCB, we explored the glucose intolerance, prediabetes, liver steatosis and insulin resistance-related changes, as well as neuroinflammation in the brain of Wistar Kyoto (WKY) rats fed a HF diet for 52 weeks. Old obese WKY rats showed robust glucose intolerance that was attenuated by palm11-PrRP31, but not by liraglutide treatment. On the contrary, liraglutide had a beneficial effect on insulin resistance parameters. Despite obesity and prediabetes, WKY rats did not develop steatosis following to HF diet feeding, even though liver lipogenesis was enhanced. Plasma triglycerides and cholesterol were not increased by HFD feeding. The liver lipid profile was significantly altered by a HF diet but remained unaffected by palm11-PrRP31 or liraglutide treatment. The HF-diet-fed WKY rats revealed astrogliosis in the brain cortex and hippocampus, which was attenuated by treatment. In conclusion, this study suggested multiple beneficial anti-obesity-related effects of palm11-PrRP31 and liraglutide in both the periphery and brain.
Mráziková L et al. Lipidized PrRP Analog Exhibits Strong Anti-Obesity and Antidiabetic Properties in Old WKY Rats with Obesity and Glucose Intolerance. Nutrients. 2023;15(2):280.
In cooperation with colleagues from the Institute for Translational Research in Biomedicine (Kaohsiung, Taiwan) we analyzed whether tissue oxidative stress in the rostral ventrolateral medulla (RVLM), where sympathetic premotor neurons reside, plays an active role in age-dependent susceptibility to hypertension in response to nitric oxide (NO) deficiency induced by systemic L-NAME treatment, and to decipher the underlying molecular mechanisms. Systemic L-NAME also evoked oxidative stress in RVLM of adult, but not young rats, accompanied by augmented enzyme activity of NADPH oxidase and reduced mitochondrial electron transport enzyme activities. The mRNA microarray analysis revealed that four genes involved in ROS production and clearance were differentially expressed in RVLM in an age-related manner. These data suggest that hypertension induced by systemic L-NAME treatment in young rats is mediated primarily by NO deficiency that occurs both in vascular smooth muscle cells and RVLM. In contrast, enhanced augmentation of oxidative stress in RVLM may contribute to the heightened susceptibility of adult rats to hypertension induced by systemic l-NAME treatment.
Chao YM, et al. Disparate Roles of Oxidative Stress in Rostral Ventrolateral Medulla in Age-Dependent Susceptibility to Hypertension Induced by Systemic l-NAME Treatment in Rats. Biomedicines. 2022;10(9):2232.
Recently, we demonstrated that chronic blockade of the renin-angiotensin system (RAS) lowered the blood pressure (BP) of adult Ren-2 transgenic rats (TGR) mainly through the attenuation of central sympathoexcitation. However, the participation of central and peripheral mechanisms in the development of high BP in immature, 6-week-old TGR males remains unclear. Chronic intraperitoneal (losartan administration (1 or 2 mg/kg/day) lowered the BP of immature TGR by reducing both sympathetic and angiotensin II-dependent BP components. Chronic intracerebroventricular administration of a lower losartan dose (1 mg/kg/day) reduced only the sympathetic BP component, whereas a higher dose (2 mg/kg/day) was required to influence the angiotensin II-dependent BP component. In conclusion, central sympathoexcitation seems to play an important role in hypertension development in immature TGR. Central sympathoexcitation is highly susceptible to inhibition by low doses of RAS-blocking agents, whereas higher doses also affect peripheral angiotensin II-dependent vasoconstriction.
Řezáčová L, et al. Both central sympathoexcitation and peripheral angiotensin II-dependent vasoconstriction contribute to hypertension development in immature heterozygous Ren-2 transgenic rats. Hypertens Res. 2022;45(3):414-423.
This study, based on the collaboration with colleagues from IOCB tried to describe the relationship between metabolic and neurodegenerative pathologies and to explore if palm11-PrRP31 could ameliorate them in obese fa/fa rat model with leptin signaling disruption. Fa/fa rats developed obesity, mild glucose intolerance, and peripheral insulin resistance but not hypertension while palm11-PrRP31 treatment neither lowered body weight nor attenuated glucose tolerance but ameliorated leptin and insulin signaling and synaptogenesis in hippocampus. We demonstrated that palm11-PrRP31 had neuroprotective features without anti-obesity and glucose lowering effects in fa/fa rats. This data suggest that this analog has the potential to exert neuroprotective effect despite of leptin signaling disturbances in this rat model.
Mráziková L, Hojná S, et al. Palmitoylated prolactin-releasing peptide treatment had neuroprotective but not anti-obesity effect in fa/fa rats with leptin signaling disturbances. Nutr Diabetes. 2022;12(1):26.
The new antidiabetic drugs, gliflozins, inhibit sodium-glucose transporter-2 in renal proximal tubules promoting glucose and sodium excretion. We examined whether these beneficial effects can also be achieved in a non-diabetic hypertensive model, namely in Ren-2 transgenic rats (TGR). Empagliflozin had no effect on plasma glucose level but partially reduced blood pressure in TGR. Although food consumption was substantially higher in empagliflozin-treated TGR compared to the untreated animals, their body weight and the amount of epididymal and perirenal fat was decreased. Empagliflozin had no effect on proteinuria, but it decreased plasma urea, attenuated renal oxidative stress and temporarily increased urinary urea excretion. Several metabolic and inflammatory parameters were also improved by empagliflozin treatment. In conclusion, empagliflozin exerted beneficial antihypertensive, anti-inflammatory and metabolic effects also in a non-diabetic hypertensive model. This study was performed in collaboration with dr. Malínská group from the Institute for Clinical and Experimental Medicine (Prague) and dr. Neckář group from our Institute.
Hojná S et al. Antihypertensive and metabolic effects of empagliflozin in Ren-2 transgenic rats, an experimental non-diabetic model of hypertension. Biomed Pharmacother. 2021;144:112246.
In collaboration with dr. Maletínská group from IOCB, we examined the impact of high-fat (HF) diet-induced obesity on potential neuroinflammation and peripheral IR in males and females of THY-Tau22 mice, a model of tau pathology. Significant sex differences in neurodegenerative signs were found in THY-Tau22. Impaired short-term spatial memory was observed in 11-month-old THY-tau22 males but not in females, which corresponds to the increased neuroinflammation colocalized with pTau(T231) in the hippocampi and amygdalae of THY-Tau22 males. A robust decrease in synaptic and postsynaptic plasticity was observed in 11-month-old females but not in males. HF diet caused peripheral but not central insulin resistance in mice of both sexes.
Kacířová M et al. Aging and high-fat diet feeding lead to peripheral insulin resistance and sex-dependent changes in brain of mouse model of tau pathology THY-Tau22. J Neuroinflammation. 2021 Jun 22;18(1):141
Salt hypertensive Dahl rats are characterized by sympathoexcitation and relative NO deficiency. We tested whether the increased blood pressure (BP) response to fasudil (Rho kinase inhibitor) in salt hypertensive Dahl rats is due to the augmented calcium sensitization in the salt-sensitive strain
and/or due to their decreased baroreflex efficiency. Our results demonstrated a substantial contribution of both factors to high BP in salt hypertensive animals. However, the BP response to the calcium channel blocker nifedipine can distinguish SS and SR rats even after calcium sensitization inhibition by fasudil, which was not seen when fasudil was administered to nifedipine-pretreated rats. Thus, the enhanced calcium entry (potentiated by sympathoexcitation) in salt hypertensive Dahl rats is the abnormality that is essential for their BP increase, which was further augmented by increased calcium sensitization in salt-sensitive Dahl rats.
Zicha J et al. Cooperation of augmented calcium sensitization and increased calcium entry contributes to high blood pressure in salt-sensitive Dahl rats. Hypertens Res. 2021;44(9):1067-1078.
In collaboration with colleagues from IOCB, we examined the neuroprotective properties of palm11-PrRP31 in double transgenic APP/PS1 mice, a model of AD-like β-amyloid (Aβ) pathology. Both liraglutide and palm11-PrRP31 treatments reduced the Aβ plaque load in the hippocampus. Palm11-PrRP31 also significantly reduced hippocampal microgliosis and reduced cortical astrocytosis, similar to the treatment with liraglutide. Moreover, Palm11-PrRP31 tended to increase neurogenesis. After the treatment with both anorexigenic compounds, we observed a significant decrease in Tau phosphorylation at Thr231, one of the first epitopes phosphorylated in AD. Both liraglutide and palm11-PrRP31 reduced the levels of caspase 3, which has multiple roles in the pathogenesis of AD. Palm11-PrRP31 increased protein levels of the pre-synaptic marker synaptophysin, suggesting that palm11-PrRP31 might help preserve synapses. These results indicate that palm11-PrRP31 has promising potential for the treatment of neurodegenerative diseases.
Holubová M et al. Liraglutide and a lipidized analog of prolactin-releasing peptide show neuroprotective effects in a mouse model of β-amyloid pathology. Neuropharmacology. 2019 Jan;144:377-387.
In cooperation with colleagues from Mossakowski Medical Research Center, Polish Academy of Sciences, we evaluated the effects of hydrochlorothiazide (diuretic) given on top of standard therapy with trandolapril and losartan (RAS blockers) and ETA receptor blockade with atrasentan in a model of chronic kidney disease. A diuretic added to the combined RAS and ETA blockade has late renoprotective effects in CKD induced by partial nephrectomy in Ren-2 transgenic rats. The diuretic treatment improved renal function (evaluated as proteinuria and creatinine clearance), renal morphology (kidney mass, glomerular volume), and histological markers of kidney damage (glomerulosclerosis index, tubulointerstitial injury).
Vaněčková I et al. Renoprotection Provided by Additional Diuretic Treatment in Partially Nephrectomized Ren-2 Transgenic Rats Subjected to the Combined RAS and ETA Blockade. Front Physiol. 2019;10:1145.
We evaluated the contribution of RAS-dependent vasoconstriction and sympathetic vasoconstriction to BP maintenance during chronic systemic or central RAS blockade in adult Ren-2 transgenic (TGR) rats. BP effect of brain RAS blockade was based solely on the reduced sympathetic vasoconstriction, while systemic RAS blockade attenuated both angiotensin II-dependent and sympathetic vasoconstriction. Surprisingly, neither peripheral nor central pressor effects of acute angiotensin II administration were enhanced in TGR compared to HanSD rats. In conclusion, sympathoinhibition represents the main mechanism of BP reduction in heterozygous TGR rats subjected to chronic central or systemic RAS blockade, while peripheral attenuation of angiotensin II-dependent vasoconstriction during systemic RAS blockade is less important.
Řezáčová L et al. Role of angiotensin II in chronic blood pressure control of heterozygous Ren-2 transgenic rats: Peripheral vasoconstriction versus central sympathoexcitation. Biomed Pharmacother. 2019 Aug;116:108996.
We investigated the hemodynamic response to acute and chronic administration of gabapentin, a ligand of auxiliary α2δ subunit of VDCCs, in adult SHR with established neurogenic hypertension. The acute gabapentin administration lowered BP and heart rate more in conscious SHR than in Wistar-Kyoto rats. Hypotensive effect of gabapentin was accompanied by a reduction of plasma norepinephrine level, depressor response to ganglionic blocker pentolinium, and low frequency component of systolic BP variability. In conclusion, besides the known L-type VDCCs involvement in the vascular effect of gabapentin, our data revealed the important role of N-type VDCCs in acute gabapentin effect on sympathetic control of BP. Gabapentin-induced changes of sympathetic nerve transmission indicated major hemodynamic mechanism of the acute response to this drug.
Behuliak M et al. Hemodynamic Response to Gabapentin in Conscious Spontaneously Hypertensive Rats. Hypertension. 2018;72(3):676-685.
We examined the potential neuroprotective effects of palm11-PrRP31, a strong anorexigenic analog with glucose-lowering properties, in THY-Tau22 mice overexpressing mutated human tau, a model of AD-like tau pathology. Treatment with palm11-PrRP31 significantly improved spatial memory. In the hippocampi of palm11-PrRP31-treated THY-Tau22 mice, tau protein phosphorylation was attenuated at Thr231, Ser396, and Ser404, the epitopes linked to AD progression. Furthermore, we observed a significant increase in the amount of postsynaptic density protein PSD95, a marker of increased synaptic plasticity. Thus, palm11-PrRP31 seems to be a potential tool for the attenuation of neurodegenerative disorders in the brain.
Popelová A et al. Novel Lipidized Analog of Prolactin-Releasing Peptide Improves Memory Impairment and Attenuates Hyperphosphorylation of Tau Protein in a Mouse Model of Tauopathy. J Alzheimers Dis. 2018;62(4):1725-1736
We compared basal and activated calcium sensitization in three forms of experimental hypertension with increased sympathetic tone and enhanced calcium entry – in spontaneously hypertensive rats (SHR), heterozygous Ren-2 transgenic rats (TGR), and salt hypertensive Dahl rats. Activated calcium sensitization was enhanced in all three hypertensive models (due to the hyperactivity of vasoconstrictor systems). In contrast, basal calcium sensitization was reduced in SHR and TGR relative to their controls, whereas it was augmented in salt-sensitive Dahl rats relative to their salt-resistant controls. Similar differences in calcium sensitization were also seen in femoral arteries of SHR and Dahl rats.
Behuliak M et al. Basal and Activated Calcium Sensitization Mediated by RhoA/Rho Kinase Pathway in Rats with Genetic and Salt Hypertension. Biomed Res Int. 2017;2017:8029728.
Although the major modes of liraglutide (glucagon-like peptide-1 receptor agonist) action are well-known, its detailed action at the metabolic level has not been studied. In collaboration with dr. Maletínská group from IOCB we explored the effect of 2-week liraglutide treatment in C57BL/6 male mice with obesity and diabetes induced by 13 weeks of high-fat diet using NMR. NMR spectroscopy identified 11 metabolites significantly affected by liraglutide treatment as compared to high-fat diet-fed control group. These metabolites included those involved in nicotinamide adenine dinucleotide metabolism, β-oxidation of fatty acids and microbiome changes, with highly interesting changes in creatinine, taurine and trigonelline, which were specific for liraglutide administration.
Bugáňová et al. The effects of liraglutide in mice with diet-induced obesity studied by metabolomics. J Endocrinol. 2017;233(1):93-104.
We analyzed the effects of three different classes of RAS blockers (losartan, captopril or aliskiren) in combination with the endothelin A receptor blocker atrasentan on blood pressure of Ren-2 transgenic rtats. All RAS blockers similarly decreased BP to normotension, their effects being mediated through substantially attenuated RAS-dependent and moderately decreased SNS-dependent vasoconstriction. Atrasentan alone partially lowered BP, while BP was normalized by combination of atrasentan with either RAS blocker. In combination therapies, BP lowering effects resulted from the attenuation of both RAS- and SNS-dependent vasoconstriction. Moreover, atrasentan-treated groups had substantially reduced NO-dependent vasodilation and significantly decreased calcium influx through L-VDCC. In conclusion, although the BP-lowering effect of combined ETA and RAS blockades in TGR is predominantly dependent on the effects exerted by RAS blockade, further effects are attributable to decreased calcium influx due to chronic ETA blockade.
Vaněčková I et al. Moderate additive effects of endothelin receptor A blockade in Ren-2 transgenic rats subjected to various types of RAS blockade. Life Sci. 2016;159:127-134.
In collaboration with colleagues from the Institute for Organic Chemistry and Biochemistry (IOCB), we published a review dealing with beneficial effects of prolactin-releasing peptide analogs in the treatment of obesity. These lipidized analogs of prolactin-releasing peptide were newly synthetized in IOCB and their cardiovascular effects were analysed in our Laboratory. Acute peripheral administration of myristoylated and palmitoylated PrRP analogs to mice and rats induced strong and long-lasting anorexigenic effects and neuronal activation in the brain areas involved in food intake regulation. Thus, the strong anorexigenic and body weight-reducing effects of palmitoylated PrRP31 and myristoylated PrRP20 make these analogs attractive candidates for antiobesity treatment.
Kuneš J et al. Prolactin-releasing peptide: a new tool for obesity treatment. J Endocrinol. 2016 Aug;230(2):R51-8. doi: 10.1530/JOE-16-0046.
Endothelin-1 is a very potent vasoconstrictor. The effects of endothelin receptor A (ETA) blockade on major vasoactive systems contributing to blood pressure regulation was evaluated in Ren-2 transgenic rats, in which hypertension is associated with the insertion of murine Ren-2 gene. We found that blood pressure lowering effect of ETA blockade was mediated by the reduced Ca influx through L-type voltage-dependent calcium channels due to missing ETA receptor-dependent vasoconstriction and attenuated angiotensin II-dependent vasoconstriction.
Vaněčková I et al: Endothelin A receptor blocker atrasentan lowers blood pressure by the reduction of nifedipine-sensitive calcium influx in Ren-2 transgenic rats fed a high-salt diet. J Hypertens. 2015, 33(1): 161-9
In cooperation with dr. Lenka Maletínská group from the Institute of Organic Chemistry and Biochemistry ASCR, where lipidized analogs of neuropeptide prolactin releasing peptide (PrRP) were synthesized, we studied the properties of these analogs in vitro (binding to receptors, stability in plasma, etc.) and their effect on food intake in rodents after peripheral administration. We found a very strong binding to the cells expressing PrRP receptor. Peripheral administration of palmitoylated PrRP to fasted mice led to long lasting reduction of food intake, which was associated with an increased neuronal activity in brain areas involved in the regulation of food intake. Long-term administration of analogs in obese mice resulted in decreased food intake, weight loss and normalization of some metabolic parameters. The results clearly showed that lipidized analogues of PrRP could become attractive candidates for the treatment of obesity.
Maletínská L et al: Novel lipidized analogs of prolactin-releasing peptide have prolonged half-lives and exert anti-obesity effects after peripheral administration, International Journal of Obesity. 2015, 39(6): 986-993.
The role of alterations in Ca2+ sensitization (RhoA/Rho kinase pathway) and Ca2+ entry via voltage-gated Ca2+ channels were studied in developing spontaneously hypertension rats. Ca2+ sensitization is attenuated and Ca2+ entry is enhanced from prehypertensive stages. Reduced expression of upstream activators of Rho kinase and lower expression of CPI-17 (endogenous inhibitor of myosin light chain phosphatase) explain changes of Ca2+ sensitization during genetic hypertension development.
Behuliak M et al: Ontogenetic changes in contribution of calcium sensitization and calcium entry to blood pressure maintenance of Wistar-Kyoto and spontaneously hypertensive rats. J Hypertens. 2015; 33(12): 2443-2454.
Enhanced production of reactive oxygen species (ROS) in the brain and/or kidney has been proposed to participate in the pathogenesis of different forms of hypertension. In our studies, we evaluated the production of superoxide radicals by plasma membrane nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase in brain and kidney of hypertensive Dahl salt-sensitive rats and transgenic rats for the murine Ren-2 renin gene. We have found increased ROS production in the kidney of both hypertensive strains, whereas no changes in ROS formation were detected in the brain.