Boris Mravec

Human adipose tissue‐derived mesenchymal stem cells expressing yeast cytosinedeaminase::uracil phosphoribosyltransferase inhibit intracerebral rat glioblastoma

Prodrug cancer gene therapy by mesenchymal stem cells (MSCs) targeted to tumors represents an attractive tool to activate prodrugs directly within the tumor mass, thus avoiding systemic toxicity. In this study, we tested the feasibility and efficacy of human adipose tissue‐derived MSCs, engineered to express the suicide gene cytosine deaminase::uracil phosphoribosyltransferase to treat intracranial rat C6 glioblastoma. Experiments were designed to simulate conditions of future clinical application for high‐grade glioblastoma therapy by direct injections of therapeutic stem cells into tumor. We demonstrated that genetically modified therapeutic stem cells still have the tumor tropism when injected to a distant intracranial site and effectively inhibited glioblastoma growth after 5‐fluorocytosine (5‐FC) therapy. Coadministration of C6 cells and therapeutic stem cells with delayed 5‐FC therapy improved the survival in a therapeutic stem cell dose‐dependent manner and induced complete tumor regression in a significant number of animals. Continuous intracerebroventricular delivery of 5‐FC using osmotic pump reduced the dose of prodrug required for the same therapeutic effect, and along with repeated administration of therapeutic stem cells increased the survival time. Intracerebral injection of therapeutic stem cells and treatment with 5‐FC did not show any detectable adverse effects. Results support the arguments to begin clinical studies for treatment of high‐grade brain tumors.

Complete regression of glioblastoma by mesenchymal stem cells mediated prodrug gene therapy simulating clinical therapeutic scenario

Suicide gene therapy mediated by mesenchymal stem cells with their ability to engraft into tumors makes these therapeutic stem cells an attractive tool to activate prodrugs directly within the tumor mass. In this study, we evaluated the therapeutic efficacy of human mesenchymal stem cells derived from bone marrow and from adipose tissue, engineered to express the suicide gene cytosine deaminase::uracil phosphoribosyltransferase to treat intracerebral rat C6 glioblastoma in a simulated clinical therapeutic scenario. Intracerebrally grown glioblastoma was treated by resection and subsequently with single or repeated intracerebral inoculations of therapeutic stem cells followed by a continuous intracerebroventricular delivery of 5‐fluorocytosine using an osmotic pump. Kaplan–Meier survival curves revealed that surgical resection of the tumor increased the survival time of the resected animals depending on the extent of surgical intervention. However, direct injections of therapeutic stem cells into the brain tissue surrounding the postoperative resection cavity led to a curative outcome in a significant number of treated animals. Moreover, the continuous supply of therapeutic stem cells into the brain with growing glioblastoma by osmotic pumps together with continuous prodrug delivery also proved to be therapeutically efficient. We assume that observed curative therapy of glioblastoma by stem cell‐mediated prodrug gene therapy might be caused by the destruction of both tumor cells and the niche where glioblastoma initiating cells reside.

Fos protein expression in mouse hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei upon osmotic stimulus: colocalization with vasopressin, oxytocin, and tyrosine hydroxylase.

The quantity and topography of activated vasopressin (AVP), oxytocin (OXY), and tyrosine hydroxylase (TH) neurons were studied immunohistochemically in the anterior, middle, and posterior portions of the PVN and SON in mice 60 min after a single injection of hypertonic saline (HS, 400 microl 1.5M, i.p.). Fos-neuropeptide double-stainings revealed: (1) Fos expression in each portion of the PVN and SON; (2) maximal number of Fos-AVP (79 cells) and Fos-OXY (50 cells) double-labelings in the middle portion of the PVN; (3) low number of Fos-TH perikarya in the PVN and their lack in the SON; (4) similar incidence (around 50%) of Fos-AVP and Fos-OXY perikarya in the SON; and (5) presence of activated AVP, OXY, and TH neurons in the periventricular, subependymal, and sub-PVN zones of the PVN. Topographic analysis revealed that the majority of AVP neurons expressing Fos occupied the dorsolateral and central part of the middle portion of the PVN. In the same PVN portion, Fos-OXY neurons occurred in similar frequency, however, they were primarily distributed along the lateral and medial margins of the PVN. In the SON, Fos-OXY cells occupied mainly its dorsal, while Fos-AVP cells predominated in its ventral part. The data clearly indicate that HS is not a selective stimulus neither for PVN nor SON itself and provide evidence that both PVN and SON AVP and OXY cells play important role in the mediation of signals induced by HS. In addition, the limited number of AVP, OXY, and TH neurons activated by HS may account for their differential functional specializations selective for stress/osmotic circuits activated by HS.

Role of nervous system in cancer aetiopathogenesis

There have been several reports on tumour tissue innervation, the effect of neurotransmitters on tumour growth, the development of metastases, and the effect of altered nervous-system activity on tumour cell proliferation. In this personal view, we summarise recent findings related to the interactions between the nervous system and tumour cells and suggest further research into the role of the nervous system in the aetiopathogenesis of cancer. Data showing the transmission of signals between the brain and tumour tissue create a complex view of the nervous system in the aetiopathogenesis of cancer. This neurobiological view of cancer aetiopathogenesis suggests that humoral and nervous pathways convey signals from tumour cells to the brain, and that the brain might consequently modulate the neuroendocrine-immune system to regulate tumour growth in peripheral tissues.

Stress‐ as Well as Suckling‐Induced Prolactin Release is Blocked by a Structural Analogue of the Putative Hypophysiotrophic Prolactin‐Releasing Factor, Salsolinol

Prolactin is secreted from the anterior lobe of the pituitary gland in response both to suckling and to stress. We recently observed that 1‐methyl‐6,7‐dihydroxy‐1,2,3,4‐tetrahydroisoquinoline (salsolinol), produced in the neurointermediate lobe of the pituitary gland, as well as in the medial basal hypothalamus, can selectively release prolactin from the anterior pituitary. Therefore, it has been proposed that salsolinol is a putative endogenous prolactin‐releasing factor (PRF). Here, we report that one structural analogue of salsolinol, 1‐methyl‐3,4‐dihydroisoquinoline (1MeDIQ), can block salsolinol‐induced release of prolactin, but does not affect prolactin release in response to thyrotropin releasing hormone (TRH), α‐methyl‐p‐tyrosine (αMpT) (an inhibitor of tyrosine hydroxylase), domperidone (a D2 dopamine receptor antagonist), or 5‐hydroxytryptophan (5‐HTP), a precursor of serotonin). 1MeDIQ profoundly inhibited suckling‐, immobilization‐, as well as formalin‐stress induced prolactin release without any influence on corticosterone secretion. The 1MeDIQ‐induced reduction in prolactin response to immobilization stress was dose‐dependent. These results suggest that salsolinol can play a pivotal role in the regulation of prolactin release induced by either physiological (suckling) or environmental (stress) stimuli.

Quantitative Evaluation of Catecholamine Enzymes Gene Expression in Adrenal Medulla and Sympathetic Ganglia of Stressed Rats

Abstract: Stress‐induced changes in mRNA levels of tyrosine hydroxylase (TH), dopamine‐β‐hydroxylase (DBH), and phenylethanolamine N‐methyltransferase (PNMT) have been expressed as relative arbitrary units compared with a control group. The aim of this study was to quantify basal and stress‐induced levels of TH, DBH, and PNMT mRNAs in rat adrenal medulla (AM) and stellate ganglia (SG) by the RT‐competitive PCR method using corresponding competitors of known concentration. In rats stressed by immobilization (IMO) once for 2 h, the concentration of mRNAs was determined in various intervals after the end of stress stimulus. In SG, the basal concentration of TH mRNA was 0.017 amol/ng of total RNA, which is approximately 30 times lower than in the AM (0.460 amol/ng RNA). The basal concentration of DBH mRNA in SG was 2.60 amol/ng of total RNA, which is about 150 times more than TH mRNA in SG but only two times less than DBH mRNA in the AM in which PNMT mRNA is present in the highest concentration. After a single 2‐h IMO, the peak elevation of TH and DBH mRNA concentration in SG occurred 24 h after the termination of stress stimulus, when their AM mRNA concentrations were already at control values. Presence of PNMT mRNA levels in the SG, of control and stressed rats has been demonstrated for the first time. Repeated IMO (7 days, 2 h daily) did not produce further increase in the mRNA concentrations compared with the elevated values found in adapted control groups. Levels of TH protein were significantly increased only after repeated IMO in SG and AM. Thus, our data show for the first time the exact concentrations of TH, DBH, and PNMT mRNA in SG and AM of rats under control and stress conditions. The lowest concentration of TH mRNA in the AM and SG supports the hypothesis that tyrosine hydroxylation is the rate‐limiting step in catecholamine biosynthesis.

The role of the vagus nerve in stroke

The initiation and progression of ischemic and hemorrhagic stroke are the result of a complex cascade of processes that determine both the extent of the lesion and long-term outcome. Several of these processes, including peripheral inflammation, neuroinflammation, and neuroplasticity are influenced by the activity of the afferent as well as efferent pathways of the vagus nerve. It was shown that vagus nerve stimulation significantly reduces the extent of stroke-induced lesion of brain parenchyma. However, the mechanisms of beneficial effect of increased vagal activity on pathological processes related to stroke remains largely unclear. The aim of this article is to describe the role of afferent and efferent vagal pathways in the mechanisms that influence the initiation of stroke as well as its detrimental effects.

Effect of rhythmic melatonin administration on clock gene expression in the suprachiasmatic nucleus and the heart of hypertensive TGR(mRen2)27 rats.

Objectives Plasma melatonin concentrations in non-dipping patients show a blunted daily rhythm. Melatonin has a capacity to improve disturbances in biological rhythms. Hypertensive TGR(mRen2)27 (TGR) rats with an upregulated renin–angiotensin system and inverted blood pressure profile were used to elucidate whether melatonin is able to influence the control of blood pressure. Design Melatonin was administered in drinking water to normotensive Sprague–Dawley (SD) and hypertensive TGR rats during the dark phase of the light: dark cycle 12: 12 for 4 weeks. Methods The effect of melatonin on blood pressure was monitored, and the expression of clock genes per2 and bmal1 and melatonin receptor MT1 in the suprachiasmatic nucleus (SCN) and the heart was measured by real time polymerase chain reaction during a 24-h cycle. Results and conclusion The administration of melatonin did not influence clock gene expression in the SCN but its effect on clock gene expression in the heart was phase dependent in both SD and TGR rats. Melatonin administration did not decrease the expression of melatonin receptors in the SCN and the heart. Melatonin did not decrease blood pressure in TGR rats but influenced the peripheral oscillator in the heart independently of the SCN. A modified function of molecular circadian oscillators in the heart can interfere with anticipation and disturb the adaptation of this organ to pressure overload.

Neurobiology of cancer: Interactions between nervous, endocrine and immune systems as a base for monitoring and modulating the tumorigenesis by the brain

The interactions between the nervous, endocrine and immune systems are studied intensively. The communication between immune and cancer cells, and multilevel and bi-directional interactions between the nervous and immune systems constitute the basis for a hypothesis assuming that the brain might monitor and modulate the processes associated with the genesis and progression of cancer. The aim of this article is to describe the data supporting this hypothesis.

You may need the vagus nerve to understand pathophysiology and to treat diseases.

Can different pathophysiological mechanisms and risk factors leading to various diseases be linked with altered transmission of signals by one common pathway? The present article provides evidence for the hypothesis that adequate vagal nerve activity reduces the risk of major diseases, via common basic mechanisms and interim risk factors. These diseases include cardiovascular disease, cancer, Alzheimers disease and the metabolic syndrome. Three basic mechanisms contribute to such illnesses: local oxidative stress and DNA damage, inflammatory reactions and excessive sympathetic responses, all of which are inhibited by vagal nerve activity. Efferent vagal activity that can be non-invasively measured by HRV (heart rate variability), derived from an ECG, is inversely related to all three basic mechanisms, to various risk factors (e.g. diabetes and dyslipidaemia) and, more broadly, to the diseases as well. Finally, vagal activity is proposed to moderate the effects of risk factors on developing such illnesses. By proposing an integrative neurobiological model of major diseases, identifying people at risk for, and treating patients with, such diseases may be done more efficiently. People with low HRV may be identified and subsequently treated by vagus nerve activation to possibly prevent or treat such illnesses. This proposed disease paradigm may have important preventative and therapeutic implications, whose clinical effects need to be investigated.