Effects of caffeic and chlorogenic acids on the rat skeletal system.

OBJECTIVE: Caffeic acid, predominantly as esters linked to quinic acid (chlorogenic acids), is a phenolic acid present at high levels in coffee. The aim of the study was to investigate effects of caffeic and chlorogenic acids on the skeletal system of female rats with normal estrogen levels and estrogen-deficient. MATERIALS AND METHODS: Caffeic acid (5 and 50 mg/kg p.o. daily) and chlorogenic acid (100 mg/kg p.o. daily) were administered for 4 weeks to non-ovariectomized and bilaterally ovariectomized mature Wistar rats, and their effects were compared with appropriate controls. Moreover, estradiol (0.2 mg/kg p.o. daily) was administered to ovariectomized rats. Bone turnover markers, mass, mineralization and mechanical properties were examined. RESULTS: Although caffeic acid at a low dose exerted some unfavorable effects on the skeletal system, at high doses, caffeic and chlorogenic acids slightly increased mineralization in the tibia and improved mechanical properties of the femoral diaphysis (compact bone). Unlike estradiol, they did not counteract the worsening of the tibial metaphysis bone strength (cancellous bone) and increases in osteocalcin concentration induced by estrogen deficiency. CONCLUSIONS: High doses of the phenolic acids slightly favorably affected the rat skeletal system independently of the estrogen status.

Platelet phospholipase A2 activity in patients with Alzheimer's disease, vascular dementia and ischemic stroke.

Phospholipase A(2) (E.C. 3.1.1.4, PLA(2)) plays an essential role in metabolism of membrane phospholipids, it is related to inflammatory reactions, secretion of amyloid precursor protein and activation of NMDA receptor after ischemia. In the present study we investigated PLA(2) activity in platelets from 37 Alzheimer's disease (AD) patients, 32 vascular dementia (VaD) patients and 32 individuals with ischemic stroke as compared to 27 healthy elderly controls. PLA(2) activity was determined using radiometric assay. Mean platelet PLA(2) activity was increased in individuals with Alzheimer's disease (p < 0.001). In VaD group the enzyme activity was between the values in AD and controls, these differences being significant from both groups. In the group of patients with ischemic stroke mean PLA(2) activity was higher either 48 h after the stroke or 7 days later (in both cases p < 0.001). The results may be particularly interesting in light of the fact, that inhibitors of PLA(2) activity are known.

Does trimetazidine exert cytoprotective activity on astrocytes subjected to hypoxia in vitro?

The aim of the present study was to establish whether trimetazidine (TMZ) is capable of protecting astrocytes against hypoxic injury. Using the model of astrocyte cell culture we tried to observe the cells treated with TMZ before, during and after hypoxia simulated in vitro. Cell viability was determined by Live/Dead (viability/cytotoxicity) Assay Kit and MTT conversion test. Apoptotic cell death was distinguished by a method using fluorescence microscopy with Hoechst 33342. The effect of the drug on the DNA synthesis was evaluated by measuring the incorporation of [3H]thymidine into DNA of astrocytes. TMZ stimulates the proliferation of astrocytes most significant one when the astrocytes are exposed to the drug in normoxia, hypoxia and/or re-oxygenation. Adding TMZ into cultures during re-oxygenation and hypoxial re-oxygenation significantly decreases the number of dead and apoptotic cells. Our experiment has proved that TMZ exerts the most significantly cytoprotective effect on astrocytes in vitro when added during hypoxia and/or re-oxygenation. We may conclude that the protective effect of TMZ depends on the sequence of drug adding and hypoxia/ re-oxygenation onset.

New analogues of bradykinin containing a conformationally restricted dipeptide fragment in their molecules.

The present paper describes the synthesis and some pharmacological properties of two new bradykinin analogues containing the ethylene-bridged dipeptide Phe-Phe in their molecules. In a further two peptides this modification was combined with acylation of the N-terminus with 1-adamantaneacetic acid. Finally, we synthesized four analogues by removing the Ser6 residue from the four peptides mentioned above. The activity of the new analogues was assayed on isolated rat uterus (RUT) and in rat blood pressure tests (BPT). The results clearly indicate that the proposed modification, alone or in combination with other changes, resulted in either a drop in antiuterotonic activity or even in conversion to an agonism. Although this tendency is not so distinct in blood pressure assays, the antagonistic potency of the new analogues is also diminished. Nevertheless, it was demonstrated that the D-amino acid in position 7 which, until recently, was considered necessary for antagonism, may be replaced, together with the amino acid occupying position 8, by a suitable, sterically restricted L,L-dipeptide unit.

Role of astrocytes in pathogenesis of ischemic brain injury.

Astrocytes play an important role in the homeostasis of the CNS both in normal conditions and after ischemic injury. The swelling of astrocytes is observed during and several seconds after brain ischemia. Then ischemia stimulates sequential morphological and biochemical changes in glia and induces its proliferation. Reactive astrocytes demonstrate stellate morphology, increased glial fibrillary acidic protein (GFAP) immunoreactivity, increased number of mitochondria as well as elevated enzymatic and non-enzymatic antioxidant activities. Astrocytes can re-uptake and metabolize glutamate and in this way they control its extracellular concentration. The ability of astrocytes to protect neurons against the toxic action of free radicals depends on their specific energy metabolism, high glutathione level, increased antioxidant enzyme activity (catalase, superoxide dismutase, glutathione peroxidase) and overexpression of antiapoptotic bcl-2 gene. Astrocytes produce cytokines (TNF-alpha, IL-1, IL-6) involved in the initiation and maintaining of immunological response in the CNS. In astrocytes, like in neurones, ischemia induces the expression of immediate early genes: c-fos, c-jun, fos B, jun B, jun D, Krox-24, NGFI-B and others. The protein products of these genes modulate the expression of different proteins, both destructive ones and those involved in the neuroprotective processes.

Effect of nebracetam on content of high-energy phosphates and morphometry of rat astrocytes in vitro. Comparison with piracetam.

The present study was initiated to examine the effect of nebracetam, a nootropic drug, on various biochemical and morphometric parameters in order to gain some insight into the mechanism of this agent action. The content of adenosine triphosphate (ATP) and phosphocreatine (PCr) and 3H-valine incorporation into proteins was measured and the morphometry was performed after nebracetam and piracetam treatment of rat astrocytes cultured in vitro with or without dibutyryl 3',5'-cyclic adenosine monophosphate (dBcAMP). Nootropics were added into the culture medium for 2 weeks at the final concentration of 10(-7) M. Cultured astrocytes treated with either nebracetam or piracetam showed decreased intracellular ATP and PCr levels. The addition of nebracetam and dBcAMP to cultures caused an increase of PCr content in astrocytes. The astrocytes treated with nebracetam showed a decrease in 3H-valine incorporation. The increase of 3H-valine incorporation into astrocytes after piracetam with dBcAMP treatment was found. Nootropic drugs change morphometric parameters (cell area, perimeter and form factor) of cultured astrocytes. It can be concluded that nootropics have differentiated influence on both the energetic metabolism and morphology of rat astrocytes in vitro.

Analogues of arginine vasopressin modified in position 2 or 3 with naphthylalanine: selective antagonists of oxytocin in-vitro.

In this study we describe the synthesis and some pharmacological properties of six new analogues of arginine vasopressin (AVP). Five of the peptides were substituted in position 2 with L-1-naphthylalanine (L-1-Nal) or D-1-naphthylalanine (D-1-Nal); one had D-1-Nal in position 3. All analogues were tested in bioassays for pressor and antidiuretic activity. We also tested the uterotonic activity of the peptides in-vitro. Two of the new peptides were moderately potent V1a and oxytocin antagonists. The modifications proposed resulted in a drop or the removal of antidiuretic activity and in the removal of pressor activity, or conversion into moderate antagonists. Two peptides ([Mpa1, (L-1-Nal)2]AVP and [Mpal, (D-1-Nal)2]AVP) which appear not to interact with V1a and V2 receptors were exceptionally selective oxytocin antagonists in vitro. These compounds with selective oxytocin antagonistic activity may be promising candidates for the development of potential tocolytic agents for the prevention of pre-term labour.

New bradykinin analogs in contraction of rat uterus.

In this study, we evaluated 20 of our previously synthesized peptides on isolated rat uterus by Holton's procedure with minor modifications, and compared their activity with that assessed previously by their ability to inhibit vasodepressor response to exogenous bradykinin (BK) in conscious rats. We used [D-Arg(0), Hyp(3), Thi(5, 8), (D-Phe)(7)]BK, the B(2) antagonist of Vavrek and Stewart as a model when designing our analogs. We observed that, in the case of the rat uterus test, the activity of peptides modified by acylation of the N-terminus with various bulky groups depends substantially on the chemical character of the substituent. We also learned that, contrary to previous examples, acylation of the N-terminus of antagonists, which contain a sterically restricted fragment in the C-terminal part, may not improve their antagonistic potencies. Besides an improved characterization of a series BK analogs, our studies have provided new information on the structure-activity relationship, which in turn may be of value in the design of more potent and selective bradykinin antagonists. The results of our studies appear to support the hypothesis of others about the presence of different subtypes of B(2) receptors in rat uterus and blood vessels.

Influence of piracetam and oxiracetam on the content of high-energy phosphates and morphometry of astrocytes in vitro.

The nootropic drugs, including piracetam (PIR) and oxiracetam (OXI) are used in the adjunctive treatment of dementia. They are thought to directly influence energetic processes in the brain and, therefore, they are supposed to improve memory and cognition. The content of adenosine triphosphate (ATP) and phosphocreatine (PCr) and 3H-valine incorporation into proteins were measured, and the morphometry was performed after PIR and OXI treatment of astrocytes cultured in vitro with or without dibutyryl 3',5'-cyclic adenosine monophosphate (dBcAMP). Nootropics were added into the culture medium for 2 weeks at a final concentration of 10(-7) M. It was shown that OXI increased ATP content in astrocytes cultured with or without dBcAMP. The increase in 3H-valine incorporation into astrocytes after PIR and OXI together with dBcAMP treatment was found. These results indicate that the presented research model allows to study energetic processes in cultured astrocytes. However, nootropic drugs changed morphometric parameters (cell area, perimeter and form factor) of cultured astrocytes as well. It can be concluded that PIR and OXI as nootropics have an opposing effect on the content of high-energy phosphates and shape of astrocytes in vitro.

Biologically active analogues of arginine vasopressin containing conformationally restricted dipeptide fragments.

In this study we described the synthesis and pharmacological properties of five new analogues of arginine vasopressin (AVP). Four of these analogues contained ethylene-bridged dipeptide Phe-Phe in positions 2 and 3; one had two N-Me-Phe residues. All new peptides were tested for vasopressor and antidiuretic activities. We also estimated the uterotonic activities of these compounds in vitro. Three analogues were highly potent V1-antagonists. One of them, namely [Cpa1,(Phe-Phe)2,3,Val4]AVP, which seemed to not interact with either V2 and oxytocic receptors, was outstandingly selective. It is interesting that the high antipressor potency of our second peptide, [(N-Me-Phe)2,3]AVP, was achieved without modification of position 1. Our results open new possibilities for the design of very potent and selective V1-antagonists of AVP.

Influence of L-naphthylalanine in position 3 of AVP and its analogues on their pharmacological properties.

We describe the synthesis and pharmacological properties of two series of analogues: one which consists of three peptides having L-1-naphthylalanine in position 3 and the second composed of analogues substituted in position 3 with L-2-naphthylalanine. All peptides were tested in bioassays for pressor and antidiuretic activities. We also checked the uterotonic activity in vitro. We observed that the activity of counterparts in both series is, in two cases, strikingly different. One of the new analogues, [(L-2-Nal)3,(D-Arg)8]VP is among the most potent antagonists of the vasopressor response to AVP. Moreover, it is the first potent V1 antagonist devoid of antiuterotonic activity. This analogue was designed without modification of position 1, which was previously thought to be essential for substantial pressor antagonism. Two other peptides, [Mpa1;(L-2-Nal)3;(D-Arg)8]VP and [Mpa1,(L-1-Nal)3,D-Arg)8]VP, are highly potent V2 agonists. The second analogue is highly selective. With the exception of [(L-2-Nal)3]AVP, which showed weak antioxytocic activity, (L-Nal)3 modification resulted in the almost complete removal of interaction of our analogues with oxytocic receptors in vitro. Our results suggest that position 3 in AVP and its analogues is important not only for binding and recognition as previously though, but also for pressor, antidiuretic and uterotonic activities. We also assume that the hindering effect caused by bulky naphthyl moiety has a significant impact on the bioactive conformations of molecules which contain Nal residue, and can thus influence their interaction with V1, V2 and oxytocic receptors.

Modification of adrenergic reactivity in rat tail artery by dietary lipids and calcium channel antagonists.

1. The antiatherosclerotic activity of dihydropyridines (DHP), potent calcium antagonists, was studied with respect to prevention of hypercontractility of perfused rat tail arteries. 2. Used for 1 month, atherogenic diet increased pressor responses to norepinephrine (NE) in Ca(2+)-free physiological salt solution (PSS), and PSS containing Ca(2+). 3. When nifedipine (NIF) or nitrendipine (NIT) was administered simultaneously with an atherogenic diet, the contractile activity of NE in Ca(2+)-free PSS was attenuated. Moreover, vasoconstrictor responses to NE in PSS containing Ca2+ were inhibited after 1-month treatment with NIT and nimodipine (NIM). 4. NIF, NIT and NIM prevented atherosclerosis-induced vascular hyperreactivity to alpha-adrenoceptor agonists in rat tail artery.

Surprising pharmacological activity of analogues designed by substitution of position 3 in arginine vasopressin (AVP) and 8-D-arginine vasopressin with L-2-napthylalanine.

In an attempt to develop more active and selective analogues of arginine vasopressin (AVP), two peptides have been designed, synthesized and tested for vasopressor (V1-receptors) and antidiuretic (V2-receptors) activities. We also estimated the uterotonic and anti-uterotonic activities of these compounds in-vitro. The first peptide, [(L-2-Nal)3] AVP is a highly active V2-agonist. The second analogue, [(L-2-Nal)3, (D-Arg)R]VP is among the most potent antagonists of the vasopressor response to AVP. Moreover, it is the first V1-antagonist devoid of anti-uterotonic activity. High antipressor potency of the second peptide was achieved without modification of position 1.

Effect of neuroleptics on phospholipase A2 activity in the brain of rats.

The effect of neuroleptics on phospholipase A2 (PLA2) activity in rat brain plasma membranes was studied. Chlorpromazine (10 mg/kg), fluphenazine (5 mg/kg), thioridazine (5 mg/kg), trifluoperazine (5 mg/kg), haloperidol (2 mg/kg), and sulpiride (100 mg/kg) were administered to rats intraperitoneally as a single dose or long-term treatment (4 weeks). The PLA2 activity was determined 24, 48, and 72 h after the last injection of a drug. The enzyme activity was decreased after a single or 4-week administration of chlorpromazine, trifluoperazine, haloperidol, and sulpiride. Fluphenazine and thioridazine caused an increase of PLA2 activity in rat brain both after a single dose and long-term administration. For the first time it was shown that neuroleptics cannot only inhibit but also increase, PLA2 activity. Elucidation of this fact requires further studies.

Effect of antidepressants on ATP content, 3H-valine incorporation and cell morphometry of astrocytes cultured from rat brain.

ATP is a potential marker of cell vialibility and growth. The content of ATP and 3H-valine incorporation into proteins were measured and the morphometry was performed after antidepressant treatment of astrocytes cultured in vitro with or without dibutyryl 3'5'-cyclic adenosine monophosphate (dB-cAMP). Antidepressants were added into the culture medium (for 24 h) at a final concentration of 10(-4)M (imipramine, amitriptyline, clomipramine, doxepine, mianserin) or 10(-5)M (maprotiline). It was shown that all antidepressants except maprotiline and imipramine increased ATP level and decreased 3H-valine incorporation into astrocytes. All drugs except clomipramine and maprotiline, diminished cell area and perimeter of astrocytes. The addition of dB-cAMP to cultures caused an increase of astrocyte form factor. It can be concluded that antidepressants have a significant effect on energy metabolism and differentiation of astrocytes cultured in vitro.

Nootropics: pharmacological properties and therapeutic use.

This review summarizes the results of studies investigating the pharmacological properties, mechanisms of action, preclinical and clinical effects of nootropic drugs, and tries to integrate this knowledge in order to identify neuronal mechanisms underlying their therapeutic benefits and side effects.

[Clinical features of lecithin-cholesterol acyltransferase deficiency]. / Obraz kliniczny deficytu acylotransferazy lecytynowo-cholesterolowej.

Lecithin-cholesterol acyltransferase (LCAT) is involved in esterify of free cholesterol and in the cholesterol esters transport from peripheral tissues to the liver. Genetically dependent lack of enzyme activity leads to Fish Eye Disease and to familial LCAT deficiency. There are specific abnormalities of plasma lipids and lipid deposits in multiple tissues (familial LCAT deficiency) or in corneal only (Fish Eye Disease). Clinical features of familial LCAT deficiency include corneal opacities, anemia, and proteinuria. Renal failure is the most frequent complication, occurring in the fourth decade. Treatment of familial LCAT deficiency is based on infusions of plasma or whole blood and on kidney transplantation.

Contractility of the tail artery in rats treated with bezafibrate and fed atherogenic diet.

Bezafibrate, a potent hypolipidemic agent, was studied as potentially preventive in the atherosclerosis-associated vascular hyperresponsiveness to alpha-adrenoceptors agonists in rats. Contractile responses to norepinephrine (NE) were determined in isolated tail arteries of rats fed an atherogenic or a standard diet. Atherogenic diet was biochemically confirmed to induce hypercholesterolemia. Used for 1 month, atherogenic diet increased pressor responses to NE in physiologic salt solution (PSS) and Ca(2+)-free PSS. When bezafibrate (100 mg/kg orally, p.o.) was administered simultaneously with atherogenic diet for 1 month, the effect was inhibited, but bezafibrate administered from day 15 of our experiment had no effect on vasoconstrictor responses to NE. Bezafibrate prevented changes in contractile responses of rat tail artery in the early stages of atherogenesis.

Behavioral effects of long-term exposure to magnetic fields in rats.

Male rats and pregnant and nonpregnant female rats of the Wistar strain were sham-exposed or exposed to static (0.49 T) or to extremely low frequency (50 Hz) magnetic fields (0.018 T) 2 h per day for 20 consecutive days. Measures of irritability, exploratory activity, and locomotion were made in that order before and after the 4th, 10th, and 17th 2-h exposures. A reliable decrease in the irritability of rats after repeated exposure to a static or undulating field was found. No significant effects of treatment conditions on open-field behavior and locomotor activity were observed. Pregnancy had no influence on the behavioral end points. These results indicate that irritability of rats may be used as a simple behavioral indicant of mammalian sensitivity to magnetic fields.

[Gangliosides in neurological pharmacotherapy]. / Gangliozydy w farmakoterapii neurologicznej.

Gangliosides take part in synaptic transmission, neuronal metabolism and development of nervous tissue. They cooperate with nerve growth factor (NGF) and have positive influence on regeneration of the nervous system impairments. There exist many behavioural and biochemical evidences of gangliosides participation in the regeneration of experimentally injured animal nervous system. The therapeutic effectivity of gangliosides in clinical practice is encouraging. Commercial preparates of gangliosides (Cronassial, Sygen) have been successfully used in the therapy of chronic neuropathies, strokes and subarachnoidal haemorrhages. Among the adverse reactions to these drugs are: local irritation, anxiety and possible detrimental effect in immunological system. Ganglioside preparations need further clinical examinations.