Temperature affects the sexual differentiation and the development of brain of tilapia, Oreochromis mossambicus. Expressed sequence tags ( EST ) derived from the developing tilapia brain had been cloned in our laboratory. In the present study, we focus on the promoter of cold-induced protein. The promoter sequence of cold-induced protein from the transcription start site ( TSS ) were cloned by 5¡¦ rapid amplification of cDNA end ( 5¡¦-RACE ) and Genome Walker DNA Walking. The bioinformatic analysis was performed on the fragment for predicting the transcription factor binding site. We used the digestion method of restriction enzymes and an electrophoresis mobility shift assay to find transcription factor binding site. The results indicated that there is a putative POU3F2 binding site in the fragment form -157 bp to -149 bp. The luciferase activity assay was performed on this site and results indicated that wild type showed the enhanced promoter activity. However, site-direct mutagenesis of this site did not result in the reduction of the promoter activity.

<p>The development of the avian embryo is dependent of heat provisioning from the parents. To increase the heat transfer to a cooled egg the Zebra finch females develop a brood patch. Mild cooling generally constricts the blood vessels but the Arterio-venous anastomoses (AVA) in the brood patch in birds dilate. This is called cold-induced vasodilation CIVD. The Zebra finches were anesthetized with isoflurane and the brood patch was stimulated with a cooling probe set at 20-21 °C. Differences in the vascular changes to cooling in broody and non- broody birds were studied by comparing males and broody females. The brood patch skin was cooled, but no cold-induced vasodilation (CIVD) was documented for the males or the broody females. Isoflurane anesthesia depresses the sympathetic nervous system activity and the results support that the mechanism for CIVD in the brood patch of Zebra finches depends on a neural pathway, but does not exclude a local non-neural mechanism.</p>

The development of the avian embryo is dependent of heat provisioning from the parents. To increase the heat transfer to a cooled egg the Zebra finch females develop a brood patch. Mild cooling generally constricts the blood vessels but the Arterio-venous anastomoses (AVA) in the brood patch in birds dilate. This is called cold-induced vasodilation CIVD. The Zebra finches were anesthetized with isoflurane and the brood patch was stimulated with a cooling probe set at 20-21 °C. Differences in the vascular changes to cooling in broody and non- broody birds were studied by comparing males and broody females. The brood patch skin was cooled, but no cold-induced vasodilation (CIVD) was documented for the males or the broody females. Isoflurane anesthesia depresses the sympathetic nervous system activity and the results support that the mechanism for CIVD in the brood patch of Zebra finches depends on a neural pathway, but does not exclude a local non-neural mechanism.

Humans are known as homeothermic endotherms. To ensure thermic balance at rest when exposed to cold, they dispose of two main thermogenic processes: shivering thermogenesis (ST) and non-shivering thermogenesis (NST). ST consists of involuntary muscle contractions and NST represents the component of Hprod that is not ST. While ST is difficult to tolerate, it is not yet known which nutrients and how much are required to stimulate NST and lower ST in the cold. Whether or not food caloric intake has an impact on the relative contribution of ST and NST to total Hprod remains to be determined. Therefore, the purpose of this thesis was 1) to quantify the effects of ingesting two quantities (1507 vs 3015 kJ) of same relative compositions on cold-induced whole-body Hprod and 2) to establish the effects of these two quantities of food on the relative contribution of ST and NST to total Hprod. Five healthy male participants were exposed to a 3h mild cold, using a liquid conditioned suit with water flowing at 15°C (COLD) or 33 °C (CON) for a total of 4 trials. Thermal, metabolic and shivering responses were measured at baseline, before and after shake ingestion. Results demonstrated that Hprod and ST intensity increased in the cold, while no significant differences were found between the ingested shakes at two different caloric equivalents. In addition, ST intensity did not change, which confirmed that NST remained the same between the two conditions. Thus, knowing that the caloric intake will not maximize the thermogenic effects in the cold (i.e. improve the comfort of the individual), is it more advantageous to bring food or additional clothing, for any activity? Clearly, more research on the exact pathways of each processes in the cold with food consumption needs to be made. To that extent, the investigation of the effect of food quality on changes in the thermogenic processes during cold exposure strikes us as a fascinating area for future research.

Smelter workers who were exposed to air-borne arsenic for a mean of 23 years, and age-matched referents, were examined with clinical, physiological, and neurophysiological methods. Exposure to arsenic in workroom air was estimated to have been around the Swedish occupational limits, which were 500 yg/m before 1975 and 50 yg/ra thereafter. An increased preval ence of Raynaud's phenomenon and a reduced finger systolic blood pressure (FSP) during local and general cooling were found in the smelter workers. Slight, but significant sub-clinical neuropathy, in the form of slightly reduced nerve conduction velocity (NCV) in two or more peripheral nerves, was more common among the arsenic workers than among the referents. There were positive correlations between cumulative exposure to arsenic, reduced NCV in three peripheral motor nerves, and decrease in FSP during cooling. Arsenic levels in urine were 1 ymole/1 (75 yg/1) in the arsenic workers and 0.1 ymole/1 in the referents. In 21 arsenic workers with no or very low exposure to vibra ting hand tools, the FSP during cooling had increased significantly after 3 years wit h the lower arsenic exposure. There was no change in FSP during the summer vacation, whereas urinary levels of arsenic decreased to normal values. Thus there seems to be a slow improvement of finger blood circ ulation which is independent of short-term fluctuations in the exposure to arsenic. No seasonal variation was found in FSP during cooling with the standardized method used. When the NCV-measurements were repeated five years later the difference between arsenic workers and referents had increased, despite the fact that 14 of the 47 arsenic workers had had no exposure to arsenic during the last 1-5 years. These observations indicate, that in subjects with long term exposure to arsenic, sub-clinical neuropathy is not reversible. Ten milligrams of Ketanserin, a serotonin receptor antagonist, was given intravenously to five arsenic workers with cold-induced vasospasm. Skin temperature and FSP during cooling increased significantly with Ketanserin as compared wit h saline solution. After oral treatment, 2 x 40 mg /day for four weeks, no significant increase of FSP during cooling or rise in skin temperature was found in six arsenic workers and eleven patients with Raynaud's phenomenon. The decrease of vasospastic tendency after intravenous injection of Ketanserin indicated that similar mechanisms might operate in arsenic-induced and other types of Raynaud's phenomenon. A general co nclusion from the five studies in this dissertation is that long-term occupational exposure to arsenic has had adverse effects on the peripheral circulation and nerve conduction. The tendency to vasospasm, but not the sub-clinical neuropathy, seemed to be reversible with decreasing exposure. / <p>S. 1-54: sammanfattning, s. 55-112: 5 uppsatser</p> / digitalisering@umu

With encouraging advancements in cell therapies, there is a requirement for an effective short-term cell preservation method, enabling time for quality assurance testing and transport to their clinical destination. This project aims to pause cells at ambient temperatures, whilst maintaining viability and function post-preservation. Ambient cell preservation bypasses ice crystal exposure and toxic solute concentrations experienced with cryogenic storage. Storage in ambient conditions also avoids use of toxic cryoprotectants and aims to greatly reduce costs and reliability on specialist machinery. Early work used HOS TE85 cells (derived from an osteosarcoma) as a model. When atmospheric factors were controlled, HOS TE85 cells demonstrated effective recovery in terms of morphology, membrane integrity (viability >90%) and fold growth expansion when paused at ambient temperature for up to 144 hours. Without atmospheric control, addition of the buffering agent HEPES (25mM) to cell medium was required to keep viability above 70%, as well as to maintain yield and continual passage following 144 hours pausing. The pausing potential of therapeutically relevant human mesenchymal stem cells (hMSCs) from three individual donors (M2, M3 and M4) was tested by keeping cells in suspension for up to 72 hours. Using standard medium with the addition of 25mM HEPES, average membrane integrity was maintained above 70%. Following pausing for between 24 72 hours, hMSC attachment efficiency, immunophenotype and tri-lineage differentiation capacity (osteogenesis, adipogenesis and chondrogenesis) remained similar to non-paused cells. Apart from a short lag phase on the first passage, hMSC fold growth expansion level was consistent with the control for all three donors over 3 x 6 day passages. The colony forming unit (CFU) efficiency of paused cells was significantly reduced when compared with non-paused M2 and M4 lines, whilst M3 retained a similar CFU efficiency to its non-paused counterpart. On return to normal culture conditions, hMSCs had comparable metabolic activity rates with non-paused cells for up to 9 hours. Stable pH is vital during pausing and additional antioxidants or apoptotic inhibiters may be required to keep average viability well-above the 70% threshold, set by the US Food and Drug Administration. Collectively, results have been encouraging and show potential for the movement towards using ambient temperature preservation as an option for the short-term storage and transport of cells for therapy.

Tauopathy is accompanied by both loss of neurons and synapses. The neuronal loss is irreversible with very low chance of functional replacement therapy. However, lost synapses could be restored with proper stimuli. Perineuronal nets (PNNs) are serving as a protecting barrier for neurons, on the other hand they are significantly decreasing the synaptic plasticity. Temporary disintegration of the PNNs by enzymatic therapy might lead to rewiring and accelerate processes of memory and learning. Model of Cold Induced plasticity leads to the withdrawal of significant number of synapses across the brain. The recovery of these could be followed in healthy and diseased animals. Moreover, it can stimulate Cold shock protein dependent neuroprotective mechanisms. This master thesis is focused on these two forms of synaptic plasticity models; forced remodeling of PNNs and model of cold induced synaptic plasticity. Both will serve as a tool to modulate processes of memory and learning in the P301S tauopathy, in mice. In detail, the work will follow changes in the number of synapses at the region of CA1 of hippocampus and synaptic protein levels at level of whole hippocampus and behavioral recovery of pre-trained long-term memory task dependent on dorsal hippocampus. Key words: Perineuronal nets, aggrecan,...

<p>In birds like the Zebra finch (Taeniopygia guttata) the female, but not the male develop a brood patch upon incubation of eggs. The brood patch functions to increase heat exchange between the bird and the eggs. Development of the brood patch includes de-feathering, increased vascularization and edema formation. The increased vascularization is due to the development of arteriovenous anastomoses, AVA. The AVA are thermoregulatory vessels involved in cold induced vasodilation, CIVD, demonstrated to occur in the brood patch. Nitric oxide, NO, which is a well known vasodilator is a candidate substance for involvement in CIVD. In this study a NO-generating gel was applied to the brood patch of male and female zebra finches. Vasodilation was found to be markedly larger in females than in males. The larger vasodilation in the female brood patch is probably because NO vasodilate AVA selectively more than any other vessels. The study also investigated whether vasodilation would cause an increase in brood patch temperature. No definite changes in brood patch temperature could be observed and no conclusions could be drawn in the matter.</p>

In birds like the Zebra finch (Taeniopygia guttata) the female, but not the male develop a brood patch upon incubation of eggs. The brood patch functions to increase heat exchange between the bird and the eggs. Development of the brood patch includes de-feathering, increased vascularization and edema formation. The increased vascularization is due to the development of arteriovenous anastomoses, AVA. The AVA are thermoregulatory vessels involved in cold induced vasodilation, CIVD, demonstrated to occur in the brood patch. Nitric oxide, NO, which is a well known vasodilator is a candidate substance for involvement in CIVD. In this study a NO-generating gel was applied to the brood patch of male and female zebra finches. Vasodilation was found to be markedly larger in females than in males. The larger vasodilation in the female brood patch is probably because NO vasodilate AVA selectively more than any other vessels. The study also investigated whether vasodilation would cause an increase in brood patch temperature. No definite changes in brood patch temperature could be observed and no conclusions could be drawn in the matter.