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Biomedical subjects

J Jakus

Publications and source records attributed to J Jakus.

At least 37 records · Page 2Linked to original sources
PubMed1996

Growth inhibitory concentrations of EGF induce p21 (WAF1/Cip1) and alter cell cycle control in squamous carcinoma cells.

Previous studies have reported inhibition of A431 squamous carcinoma cell growth by nanomolar concentrations of epidermal growth factor (EGF), a potent mitogen for cells of epithelial origin. In this study, we examined potential mechanisms through which inhibition of keratinocyte growth mediated by EGF might occur by analysing components of the cell cycle regulatory machinery in A431, HN6 and HN30 keratinocytes in the presence of growth inhibitory or growth stimulatory doses of EGF. Treatment of cells with 25 pM EGF produced an increase in [3H]thymidine incorporation in A431, HN6 and HN30 cells, with respect to control cultures. Exposure to 2.5 nM EGF reduced [3H]thymidine incorporation in A431 cells and HN6 cells to 11% and 70% of control levels, respectively, whereas HN30 cells continued to proliferate in the presence of EGF. [3H]thymidine incorporation assays carried out over 24 h revealed repression of DNA synthesis in A431 cells after 12 h exposure to 2.5 nM EGF compared to untreated cells. Flow cytometry studies demonstrated accumulation of cells in G0/G1 after addition of 2.5 nM, but not 25 pM EGF. Western blot analysis revealed elevation of p21 (WAF1/CIP1/SDI1) protein levels in A431 and HN6 cells under growth-inhibitory conditions. Stimulatory doses of EGF did not induce p21 in these cells. Northern blot hybridization demonstrated elevated levels of p21 mRNA within 4 h of exposure of A431 cells to 2.5 nM EGF, which remained elevated above basal levels at 24 h. In vitro kinase assays demonstrated temporal differences in CDK2 and CDK6 activities which were related to EGF concentration. Immunocomplex Western blotting demonstrated increased association of p21 with CDK2 and CDK6 in A431 cells treated with 2.5 nm EGF. Furthermore, temporal alterations in the association of PCNA with p21 and with CDK6 were observed. The data indicate that p21 is a likely mediator of EGF-induced growth-inhibition, probably through mechanisms involving sequestration of PCNA and inhibition of CDK activity.

Base Sequence↗
PubMed1996

Synaptosomal Na, K-ATPase during forebrain ischemia in Mongolian gerbils.

We studied the activity and kinetic parameters of synaptosomal Na, K-ATPase during 15 min of forebrain ischemia and following 60 min of reperfusion produced by reversible common carotid occlusion in Mongolian gerbils. A synaptosomal fraction was obtained by both differential centrifugation of brain tissue homogenate and centrifugation of crude mitochondrial fraction at a discontinual sucrose density gradient. We found two components of ATP concentration dependence of ATP hydrolysis that represent two types of ATP-binding sites: high affinity and low affinity. Neither ischemia nor reperfusion affected kinetic parameters of a high-affinity site. However, low-affinity site parameters were affected by both ischemia and ischemia followed by reperfusion. Maximal velocity (Vmax) decreased by 43 and 42% after ischemia and after ischemia/reperfusion, respectively. The apparent Km for ATP decreased by 52% after ischemia and by 47% after ischemia/reperfusion. The apparent affinities for K+ and Na+ were determined from the ATP hydrolysis rate as a function of Na+ and K+ concentrations. We found the half-maximal activation constant for K+ (KaK+) increased by 60% after ischemia and by 146% after ischemia/reperfusion. On the other hand, we found that KaNa+ decreased significantly after ischemia/reperfusion (16%). We concluded that it is the dephosphorylation step of the ATPase reaction cycle that is primarily affected by both ischemia and ischemia/reperfusion. This might be caused by alteration of the protein molecule and/or its surroundings subsequent to ischemia.

Animals↗
PubMed1996

Laryngeal patency and expiration reflex following focal cold block of the medulla in the cat.

The involvement of the intermediate area and Bötzinger complex (BOT) of the rostral ventral respiratory group (r-VRG) in laryngeal control and generation of the expiration reflex were studied in anaesthetized non-paralyzed cats Focal cooling (to 20 degrees C) of the nucleus paraambigualis (NPA) caused changes in the frequency and timing of breathing with the concomitant rise in laryngeal resistance. Cooling of the nucleus ambiguus resulted in a consistent drop in laryngeal resistance. Alterations in timing and intensity of breathing but no changes in laryngeal patency were found during cooling of the BOT. The expiration reflex was inhibited by cooling of either the NPA or BOT. The role of these medullary regions in the control of laryngeal patency and central integration of the expiration reflex is discussed.

Airway Resistance↗
PubMed1995

Multifunctional ventral respiratory group: bulbospinal expiratory neurons play a role in pudendal discharge during vomiting.

Pudendal motoneurons are activated in phasic bursts during the retching and expulsion phases of vomiting. The resulting contraction of the anal and urethral sphincters serves to maintain continence during the large increase in abdominal pressure that occurs during vomiting. We evaluated the contribution of bulbospinal expiratory neurons located in the portion of the ventral respiratory group (VRG) caudal to the obex (nucleus retroambigualis) to the control of pudendal motoneurons during fictive vomiting in decerebrate, paralyzed cats. Pudendal nerve discharge is abolished by cutting the axons of caudal VRG expiratory neurons as they cross the midline between the obex and C1 before descending in the spinal cord. All caudal VRG expiratory neurons that were antidromically activated from the sacral spinal cord, where the pudendal motor pool (nucleus of Onuf) is located, discharged strongly during the end of the expulsion phase of vomiting. However, only a small proportion of these neurons was active in phase with pudendal discharge during the retching phase. The apparent involvement of caudal VRG expiratory neurons in the control of pudendal motoneurons during vomiting is another example of the multifunctional role that can be played by respiratory-related neurons in the mammalian nervous system.

Animals↗
PubMed1995

Identification of a substrate site for transglutaminases on the human protein synthesis initiation factor 5A.

Protein synthesis initiation factor 5A (eIF-5A) from human erythrocytes was found to be a substrate for both plasma transglutaminase (Factor XIIIa) and guinea pig liver transglutaminase (GPLTG). When purified eIF-5A was incubated with GPLTG or Factor XIIIa in the presence of succinylated beta-casein, a covalent complex was identified. By isolating and analysing the product of the transglutaminases (TGases) reaction, the site of modification on eIF-5A has been identified as the unique amino acid hypusine. The complex beta-casein.eIF-5A was enzymatically digested with proteinases and the predicted covalent cross-link of gamma-glutamyl-omega-hypusine was isolated from the digests by ion-exchange chromatography and purified by reversed-phase h.p.l.c. Acid hydrolysis of the purified dipeptide yielded equimolar amounts of hypusine and glutamic acid. Furthermore, fast atom bombardment m.s. analysis confirmed the isomer assignment to be gamma-glutamyl-omega-hypusine. These data indicate that hypusine-50 of the eIF-5A chain functions as acyl acceptor substrate for TGases, and reveal that eIF-5A may be cross-linked to intracellular proteins by TGases. Because the precise function of eIF-5A is still unknown, our results appear particularly stimulating in the light of the recent finding of a new biological role for this protein as a cellular factor binding specifically to the human immunodeficiency virus-1 Rev activation domain [Ruhl, Himmelspach, Bahr, Hammerschmid, Jaksche, Wolff, Auschauer, Farrington, Probst, Bevec and Hauber (1993) J. Cell Biol. 123, 1309-1320].

Animals↗
PubMed1994

Brain areas essential or non-essential for emesis.

This study was undertaken to better delineate those brain regions that are either essential or non-essential for vomiting. Fictive vomiting, identified by a characteristic pattern of respiratory nerve discharge, was induced by a combination of emetic drugs and electrical stimulation of abdominal vagal afferents in decerebrate, paralyzed cats. Regions non-essential for coordinating vomiting included the entire cerebellum, structures rostral to the medullary retrofacial nucleus, and spinal cord. Fictive coughing was also elicited following cerebellar removal but was not studied after other procedures. The respiratory-related components of fictive vomiting were abolished by large lesions or kainic acid injections in the lateral medulla at the level of the retrofacial nucleus, where respiratory pre-motor and motor neurons are known to exist. Electrical stimulation of this region of the brainstem failed to evoke vomiting. The results of the present study are consistent with our previous electrical stimulation [Brain Res., 270 (1983) 154-158] and c-fos [J. Neurosci., 14 (1994) 871-888] studies and the hypothesis that emesis is coordinated not by a unique, well-defined 'vomiting center' but rather by a distributed control system located in the medulla between the levels of the obex and the retrofacial nucleus.

Abdominal Muscles↗
PubMed1994

Plasticity of emesis to a 5-HT3 agonist: effect of order of visceral nerve cuts.

Serotonin type 3 receptors are critical for vomiting induced by radiation and cancer chemotherapy. To investigate the mechanisms involved, we analyzed the effects of abdominal visceral denervation using cats. Thresholds for vomiting induced by the serotonin-3 receptor agonist phenylbiguanide varied depending on the order of chronic nerve transections. Lower thoracic vagotomy, splanchnicectomy, and their sequential combination increased thresholds. However, when splanchnicectomy, which causes a large increase in threshold, was followed 14-51 days later by vagotomy, thresholds dropped by an average of 40%. These results demonstrate plasticity in, and potential for modulation of, the emetic reflex.

Animals↗
PubMed1994

Organization of vestibular inputs to nucleus tractus solitarius and adjacent structures in cat brain stem.

The vestibular system is involved in maintaining stable blood pressure and respiration during changes in posture and is essential for eliciting motion sickness-related vomiting. Because the nucleus tractus solitarius (NTS) participates in the regulation of sympathetic and inspiratory outflow and the triggering of emesis, we tested the hypothesis that this region receives vestibular inputs in cats. In one set of experiments, microinjections of the tracer Phaseolus vulgaris leucoagglutinin into the medial and inferior vestibular nuclei labeled projections to the middle and lateral regions of the NTS. In electrophysiological experiments, electrical stimulation of the vestibular nerve modified the firing rates of neurons located in the same regions. Some neurons with vestibular inputs received convergent signals from the abdominal vagus nerve and could potentially mediate motion sickness-related vomiting. Others received convergent baroreceptor inputs and could act as a substrate for some components of vestibulosympathetic reflexes. In contrast, inspiratory neurons in the dorsal respiratory group received little vestibular input, suggesting that vestibulorespiratory reflexes are mediated by cells located elsewhere.

Animals↗
PubMed1993

Vestibular effects on respiratory outflow in the decerebrate cat.

Recordings were made from phrenic, abdominal and intercostal nerves following electrical stimulation of the vestibular nerve to test for the presence of vestibulo-respiratory connections in cats that were decerebrate, paralyzed, and artificially ventilated. Short stimulus trains (2 or 3 shocks) at intensities < or = 125 microA typically elicited responses bilaterally in all of the respiratory nerves; the onset latency of the evoked activity was < 15 ms from the effective shock. The mean peak-to-peak amplitudes of integrated vestibulo-respiratory responses were 15% of the average amplitude of spontaneous respiratory-related discharges in the case of the phrenic nerve and 100% in the case of the abdominal nerve. The vestibulo-respiratory reflexes, as well as vestibulo-sympathetic responses recorded from the splanchnic nerve, could be abolished by injections of the excitotoxin kainic acid confined primarily to the medial and adjacent inferior vestibular nuclei. The physiological role of vestibulo-respiratory connections is yet to be determined, but possible functions include adjustments of respiration during changes in posture, assistance in venous return to the heart during movements that might lead to orthostatic hypotension, and direct participation in the execution of specific movements and the maintenance of some postures.

Animals↗
PubMed1993

Features of the spermidine-binding site of deoxyhypusine synthase as derived from inhibition studies. Effective inhibition by bis- and mono-guanylated diamines and polyamines.

Several types of basic compounds structurally related to spermidine, one of the substrates for deoxyhypusine synthase, were tested as inhibitors of this enzyme. The results indicate that inhibitory compounds associate with the enzyme at the site of spermidine binding and must possess two charged primary amino or guanidino groups, or one of each. The efficiency of inhibition is related to the maximum possible distance between the primary amino groups and is adversely affected by substitutions on the secondary amino group or in the carbon chains of polyamines. The mono-guanyl derivatives are much more effective inhibitors than the parent amines or their bis-guanylated counterparts, N1-guanyl-1,7-diaminoheptane being the most effective compound with a Ki value of about 10 nM. Based on these observations we have proposed a model for the spermidine-binding site of deoxyhypusine synthase. Studies with Chinese hamster ovary cells reveal a direct correlation between prevention of hypusine formation by several guanyldiamines and their in vitro inhibition of deoxyhypusine synthase. This evidence for disruption of the initial step in the post-translational maturation of eukaryotic initiation factor 5A provides a basis for the potential control of protein biosynthesis and cell proliferation.

Animals↗
PubMed1992

Characteristics of augmented breaths provoked by almitrine bismesylate in cats.

The contribution of almitrine bismesylate to the occurrence and pattern of augmented breaths was studied in fifteen spontaneously breathing, anaesthetized cats. Breathing was via a tracheostomy, while the laryngeal resistance to airflow was measured with the larynx isolated in situ. Almitrine bismesylate at a dose of 0.5 mg kg-1 of body weight was injected intravenously in the intact animals and following bilateral vagotomy which spared the right recurrent laryngeal nerve. Almitrine injected intravenously elicited augmented breaths within the first 45 s in thirteen cats and within 1 min in the remaining two cats. During augmented breaths inspiratory and expiratory airflows rose, the mean increases being 385.2 and 159.6% respectively above the controls (P less than 0.01). The inspiratory laryngeal resistance declined to 77.7% of the control (P less than 0.01) and expiratory laryngeal resistance increased by 95.4% above the control level (P less than 0.01). The inspiratory and expiratory times were prolonged by 56 and 58% compared with baseline breathing. Following the augmented breaths the respiratory airflows exceeded baseline values, the respiratory timing was slightly reduced, and the inspiratory laryngeal resistance was significantly lowered below the control level (P less than 0.01). The expiratory laryngeal resistance showed the same trend without statistical significance. Bilateral vagotomy abolished the occurrence of augmented breaths following almitrine injection.

Airway Resistance↗
PubMed1990

Pattern of breathing and laryngeal patency following almitrine bismesylate in anesthetized cats.

The effects of almitrine bismesylate on the pattern of breathing and laryngeal caliber were investigated in anesthetized, spontaneously breathing cats. Breathing occurred via a tracheostomy, while the laryngeal resistance to airflow was measured with the larynx isolated in situ. Almitrine bismesylate in a dose of 0.5 mg/kg of body weight was injected intravenously three times: in the intact animals, following bilateral vagotomy (sparing the right recurrent laryngeal nerve), and after a sensory denervation of the larynx. In each condition, almitrine stimulated ventilation by significantly increasing inspiratory airflow, by increasing the respiratory rate in the 10th and the 20th minutes following injection, and by reducing laryngeal resistance to airflow in both the inspiratory and expiratory phases. The stimulatory effects of almitrine were most apparent in the intact animals. Following vagotomy, these significant changes in the pattern of breathing as well as the enlarged laryngeal caliber persisted. However, these latter effects were short-lived and were followed by narrowing of the laryngeal lumen. Laryngeal afferents were not essential for the response to almitrine.

Airway Resistance↗
PubMed1990

Effects of focal cooling of medulla oblongata structures on quiet breathing in cats.

Experiments were carried out on 16 anaesthetized, non-paralysed cats to determine the effects of unilateral, successive focal cooling of the nuclei of the dorsal and ventral respiratory groups (DRG, VRG) of the medulla oblongata on quiet breathing parameters. The results of cold block tests of the respiratory nuclei showed that: 1. Compared with the control state, cooling of the ventrolateral part of the nucleus solitarii (vl. NTS) and the rostral part of the nucleus retroambigualis (r. NRA) to 20 degrees C or 15 degrees C decreased the respiration rate (p less than 0.001), prolonged the inspiration time (p less than 0.01 and p less than 0.001 respectively) and the development of apneustic breathing. A decrease in the inspiratory pleural pressure values (p less than 0.01) was found after cooling the r. NRA region to 15 degrees C. In 45% of the cases of cooling of the vl. NTS and 66.7% of cooling of the r. NRA to 15 degrees C, an incidence of short inspiratory efforts was observed. 2. Focal cooling of the nucleus retrofacialis (nucl. RF) region to 20 degrees C always arrested rhythmic respiration. 3. The effects of unilateral focal cooling of the respiratory nuclei were always bilaterally symmetrical and, after discontinuing cooling, reversible. 4. The findings indicate that the inspiratory neurones of the r. NRA participate more in regulation of the intensity of inspiration than those of the vl. NTS, while the nucl. RF region may be a part of central regulatory mechanisms essential for the maintenance of rhythmic breathing in cats.

Animals↗
PubMed1987

Respiration and airway reflexes after transversal brain stem lesions in cats.

The effect of brain stem transection at different levels of the pons Varolii and the medulla oblongata on respiration and on cough and the aspiration and expiration reflex elicited by mechanical stimulation of the relevant parts of the respiratory tract was studied in experiments on 13 anaesthetized, unparalyzed cats. The results of 142 respiratory reflex elicitation tests showed that: 1. Compared with the control state, transection of the upper and middle part of the pons Varolii and transection at the level of the pontomedullary junction reduced the respiration rate (p less than 0.001), increased the duration of inspiration and expiration (p less than 0.001, transection 10 mm rostrally to the obex) and gave rise to apneustic breathing (8 mm), or to tonic, respiration-modulated activity of the phrenic nerve and diaphragm (6 mm). 2. Successive transection of the pons and the pontomedullary junction region led chiefly to a drop in maximum expiratory pleural pressure values (p less than 0.01-0.001) during cough and the expiration reflex and to a drop in maximum inspiratory pleural pressure values during the aspiration reflex (p less than 0.02-0.001). 3. Transection of the upper part of the medulla oblongata always led to permanent arrest of rhythmic respiration, during which cough and the expiration reflex could not be elicited while the aspiration reflex persisted (though in a weakened form). This state was followed by gasping, during which only a highly elicitable aspiration reflex persisted. 4. It can be assumed from the above findings that the central mechanisms responsible for the development of powerful expiratory efforts in cough and the expiration reflex could be localized in the pons Varolii, while those integrating the aspiration reflex are probably localized mainly in the medulla oblongata.

Animals↗
PubMed1987

Bulbar respiratory activity during defensive airways reflexes in cats.

Experiments in anaesthetized nonparalyzed cats indicated that mechanical stimulation of the airways and a longitudinal split of the brainstem cause marked alterations of the regulatory function of the bulbar respiratory neurones. The resulting changes in breathing and in defensive airways reflexes depend on intactness of the relevant structures of both halves of the "respiratory centre" and their interconnections.

Action Potentials↗