Effect of HBO therapy on adipose-derived stem cells, fibroblasts and co-cultures: In vitro study of oxidative stress, angiogenic potential and production of pro-inflammatory growth factors in co-cultures1.

BACKGROUND: A key moderator of wound healing is oxygen. Wound healing is a dynamic and carefully orchestrated process involving blood cells, cytokines, parenchymal cells (i.e. fibroblasts and mesenchymal stem cells) and extracellular matrix reorganization. Human adipose derived stem cells as well as human fibroblasts produce soluble factors, exhibit diverse effects on inflammation and anti inflammation response and are involved in wound healing processes.Hyperbaric oxygen therapy is an effective adjunct treatment for ischemic disorders such as chronic infection or chronic wounds. In vitro effects of hyperbaric oxygen therapy on human cells were presented in many studies except for those on mono- and co-cultures of human adipose derived stem cells and fibroblasts. OBJECTIVE: The aim of this study was to investigate the effects of hyperbaric oxygen therapy on mono- and co-cultures of human adipose derived stem cells and fibroblasts. METHODS: Mono- and co-cultures from human adipose derived stem cells and fibroblasts were established. These cultures were exposed to hyperbaric oxygen therapy every 24 h for five consecutive days. Measuring experiments were performed on the first, third and fifth day. Therapy effects on the expression of VEGF, IL 6 and reactive oxygen species were investigated. RESULTS: After exposure to hyperbaric oxygen, cell culturess showed a significant increase in the expression of VEGF after 3 and 5 days. All cultures showed significantly reduced formation of reactive oxygen species throughout the experiments. The expression of IL-6 decreased during the experiment in mono-cultures of human adipose derived stem cells and co-cultures. In contrast, mono-cultures of human skin fibroblasts showed an overall significantly increased expression of IL-6. CONCLUSIONS: Hyperbaric oxygen therapy leads to immunmodulatory and proangiogenetic effects in a wound-like enviroment of adipose derived stem cells and fibroblasts.

A new approach for fat removal in a discontinuous autotransfusion device-concept and evaluation.

BACKGROUND: Fat present during blood salvage in orthopaedic or cardiac surgery can pose a risk of fat embolism and should be eliminated before transfusion. Based on observations of central fat accumulation at the bottom of Latham bowls, a fat reduction program was developed using two volume displacements, where blood temporarily is removed and respun in the bowl to force the fat through the RBC sediment. MATERIALS AND METHODS: Pooled ABO-matched RBC and FFP were adjusted to a haematocrit of 10%, and human fat tissue added to a concentration of 1·25 vol%. In six experiments, blood was processed with the new-generation cell salvage device CS Elite in a newly developed fat reduction program in bowls of three sizes. Volumetric quantification of fat was performed after centrifugation of blood samples in Pasteur pipettes. From volumes, haematocrits and the concentrations of fat, RBC recovery and fat elimination rates were calculated. RESULTS: Fat removal rates of 93·2 ± 2·8, 97·0 ± 2·1 and 99·6 ± 0·3% were observed with a 70-ml, 125-ml and 225-ml bowl, respectively, and even higher rates when removal rates were calculated one cycle. At the same time, high RBC recovery and plasma elimination rates were maintained, not significantly different to the default program mode. CONCLUSION: Modifications in process parameters and sequence led to a fat reduction program that significantly improves fat removal with the Cell Saver Elite from 77·4 ± 5·1% in the default mode to an average of 98·6 ± 1·1%, yielding results equivalent to the continuous cell salvage system (C.A.T.S).

Performance of a new-generation continuous autotransfusion device including fat removal and consequences for quality controls.

BACKGROUND AND OBJECTIVES: Cell salvage plays a key role in blood conservation. To maintain high performance, quality management is recommended. Accordingly, a new-generation autotransfusion device was tested for its performance and compared with its predecessor. Two different calculations of quality parameters were applied. MATERIALS AND METHODS: In an experimental study, the continuous autotransfusion devices CATSmart and Continuous Autotransfusion System (C.A.T.S) plus were tested using banked blood adjusted to a haematocrit of 20% and anticoagulated with heparin 5 U/L. Test blood was processed using an emergency programme, a high-quality programme/smart wash programme and a low-volume wash programme. Samples were taken after the production of 200 mL of red blood cells (RBC) and after the final emptying of the separation chamber. In an additional set of tests, blood containing 1·25% fat was processed with both devices to examine fat removal. RESULTS: Both devices demonstrated an equally high performance with regards to product hematocrit (Hct); RBC recovery; and elimination rates of protein, heparin and fat. The high fat elimination rate (>99·8%) reported for C.A.T.S plus was confirmed for CATSmart, regardless of the used programme. Samples taken during the ongoing process show a higher haematocrit and RBC recovery rate than samples taken after the final emptying of the separation chamber. Interface sensors were not affected by fat in the blood. CONCLUSIONS: The new-generation autotransfusion device CATSmart is not inferior to its predecessor and shows high performance with regards to RBC recovery, plasma and fat elimination in all programme modes. Samples for quality controls should be taken during blood processing.

Effects of peripherally and centrally applied ghrelin in the pathogenesis of ischemia-reperfusion induced injury of the small intestine.

Ghrelin is an important hormone involved in the control of the human appetite center. Recently, protective properties of this hormone have been recognized in various models of impairment of the gastric mucosa, including stress, ischemia and reperfusion (I/R). Ghrelin is predominantly secreted by the gastric mucosa of stomach, but there are other sources of ghrelin, for example in the hypothalamus and various parts of the central nervous system (CNS) that should be taken into consideration. This hormone exerts biological effects via the activation of growth hormone secretagogue receptor (GHSR), the presence of which was confirmed in different parts of the gastrointestinal (GI) tract and midbrain structures. Although substantial evidence of the divergent biological effects of ghrelin and the mechanism of its action has been emphasized, the precise mechanisms of ghrelin which affords GI protection is still unclear. Particularly, there is a sparse amount of evidence concerning its action on the GI system. The major aim of the present study was to evaluate the importance of peripherally and centrally administered ghrelin at different times of the ischemia and reperfusion (I/R period in the modulation of resistance of the intestinal mucosa to the injury induced by ischemia and subsequent reperfusion. Secondly, we wanted to evaluate the possible mechanism of the action of ghrelin with a particular focus on its influence on the intestinal blood flow. Male Wistar rats were divided into 4 series (A-D) of the experimental groups (n=7). In series A the importance of peripherally administered ghrelin at different time of I/R period was studied. In series B the importance of centrally administered ghrelin at different time of I/R period was evaluated. In series C and D, the mechanisms of peripherally and centrally administered hormone were examined, respectively. Two models of the I/R period were selected: short lasting (30/60 min) and long lasting (60/120 min). The following drugs were used: ghrelin (50 µg/kg i.p. or 1 nmol in 10 µl i.c.v.), 6 hydroxy dopamine (50 mg/kg i.p.), nadolol (0.5 mg/kg i.p.), calcitonin gene related peptide fragment (CGRP(8-37), 100 µg /kg i.p.), capsaicin (5-10 mg/100 ml solution s.c.). The mesenteric blood flow (MBF-ml/min), the intestinal microcirculatory blood flow (LDBF-PU), the arterio-venous oxygen difference (AVO(2)-ml/O(2)/100 ml blood), and the intestinal oxygen uptake (VO(2)) in ml O(2)/min were measured. Mucosal impairment was assessed planimetrically with the use of a digital photo analyzer (LA) and histologically with the use of the six-point Park/Chiu scale. Peripheral administration of ghrelin evoked marked increase of MBF and LDBF by 42% and 48%, respectively, with significant reduction of LA by 38%. When ghrelin was administered at the beginning of the reperfusion period during the short I/R period or prior to the long lasting I/R period, the vascular reactions and protective effects were reduced, but not completely abolished. The central administration of ghrelin before the short I/R period significantly increased the MBF and LDBF by about 32% and 35%, respectively, as well as LA reduction by about 20% in comparison to the control group. However, when ghrelin was administered prior to the long I/R period or after the onset of completed ischemia, neither vascular nor protective effects were noticed. Sensory denervation and the blockade of the CGRP1 receptors totally blocked the protective and hyperemic effects of the peripherally administered ghrelin. Selective blockade of the adrenergic system or blunting of the vagal nerves (vagotomy) significantly but not totally eliminated the effects of centrally applied ghrelin, which were abolished when both adrenergic and parasympathetic pathways were ablated. These results indicate that ghrelin applied centrally or peripherally markedly increases resistance of the intestinal tissue during the I/R period induced mucosal and hyperemic impairment evoked by I/R. Ghrelin is an important mediator of the increase in the intestinal microcirculation and elevation of the intestinal metabolism, which seems to be, at least in part, responsible for the observed protection of the intestine subjected to I/R. Impairment of this microvasculature response due to I/R seems to be responsible for a markedly observed weaker effect of ghrelin when this hormone was administered after the ischemic period. The lack of a protective effect observed after central administration of this peptide against a long lasting I/R period is probably due to damage of neural pathways caused by I/R. Finally, the peripheral activity of ghrelin in the intestine is mediated by the sensory neurons with a prominent role of CGRP released from their endings. However, this peripheral action of ghrelin depends upon the proper functioning of both the sympathetic and parasympathetic system.

Mucosal strengthening activity of central and peripheral melatonin in the mechanism of gastric defense.

This review summarizes the involvement of centrally and peripherally applied melatonin, a major hormone of pineal gland, in the mechanism of gastric mucosal integrity, gastroprotection and ulcer healing. Melatonin was originally shown to attenuate gastric mucosal lesions but the controversy exists in the literature as to whether melatonin derived from the pineal gland, considered as the major source of this indole or rather that locally generated from L-tryptophan within gastric mucosa, plays predominant role in the mechanism of gastrointestinal integrity. Both, intragastric (i.g.) and intracerebroventricular (i.c.v.) administration of melatonin and its precursor, L-tryptophan to rats without or with removed pineal gland by pinealectomy attenuates in the dose-dependent manner the formation of on gastric lesions induced by topical irritants and water immersion restraint stress (WRS). Melatonin accelerated the gastric ulcer healing and this was accompanied by the rise in gastric blood flow (GBF), the plasma melatonin and gastrin levels, the mucosal generation of PGE(2) and luminal NO content. Pinealectomy, which suppresses the plasma melatonin levels, markedly aggravated the gastric lesions induced by WRS. Concurrent supplementation of pinealectomized animals with melatonin or L-tryptophan, the melatonin precursor, attenuated the lesions induced by WRS. Treatment with luzindole, an antagonist of Mel(2) receptors, or with L-NNA, the NO-synthase inhibitor, significantly attenuated melatonin- and L-tryptophan-induced protection and the acceleration of ulcer healing and the accompanying increase in the GBF and luminal content of NO. We conclude that 1) exogenous melatonin and that released from the L-tryptophan attenuate lesions induced by topical irritant such as ethanol and WRS via interaction with MT(2) receptors and due to an enhancement of gastric microcirculation, probably mediated by NO and PG derived from cNOS, iNOS and COX-2 overexpression and activity, and 2) the pineal gland plays an important role in the limitation of WRS-induced gastric lesions and acceleration of ulcer healing via releasing melatonin predominately at night time, that exerts gastroprotective and ulcer healing actions.

[Autologous transfusion -- from euphoria to reason: clinical practice based on scientific knowledge (Part II). Intraoperative blood salvage with blood irradiation -- from an anaesthesiological point of view]. / 3. Internationales Symposium: "Autologe Transfusion -- Von der Euphorie zur Ratio: Praktisches Handeln aus wissenschaftlicher Sicht" (Teil II).

The requirement for intraoperative blood salvage (IBS) in cancer surgery stems from the high transfusion rate, the unfavourable effects of an anaemia, and the impact of transfusion risks like immunomodulation in tumor patients. The advantages of IBS are availability, the low waste rate, and the excellent quality of this autologous, unstored blood. The only effective elimination of the risk of tumor cell dissemination after retransfusion of wound blood is achieved by blood irradiation. The combination of the established methods of IBS and blood irradiation is practical, and allows a very efficient saving of blood. For Jehovah's witnesses it may open the possibility for tumor surgery. From an anaesthesiological point of view it is part of the therapy, but any context that puts it compatible to medicolegal regulations is welcome. For him as the one responsible for the therapy of intraoperative blood loss it represents the safest and best blood for an optimal hemotherapy in tumor patients. In addition, first data indicate a better outcome of these patients.

Involvement of central and peripheral histamine H(3) receptors in the control of the vascular tone and oxygen uptake in the mesenteric circulation of the rat.

Data concerning cardiovascular effects of peripherally and centrally located histamine H(3) receptor stimulation are contradictory, and despite excessive studies their role in the control of the cardiovascular function have not been cleared yet. Effect of histamine H(3) receptors activation have been attributed to modulation of sympathetic system activity but exact role of peripherally and centrally located histamine H(3) receptors stimulation in the modulation of vascular tone of the mesentery and intestinal metabolism remains unexplored. Aim of the present study was to evaluate the role of centrally and peripherally located histamine H(3) receptors in the modulation of vascular tone of the mesentery and metabolic activity of intestinal tissue. In anesthetized rats total mesenteric blood flow (MBF), mucosal intestinal blood flow (LDBF), intestinal oxygen uptake (VO(2)) and arterial pressure (AP) were determined. Intestinal arterial conductance (C) was also calculated. Administration of the selective histamine H(3) receptor agonist imetit (10 micromol/kg i.a) evoked marked changes in hemodynamic and metabolic parameters; MBF, LDBF, C and VO(2) were significantly increased, whereas AP was significantly decreased. Pretreatment with histamine H(3) receptor antagonist clobenpropit (4 micromol/kg i.a.) abolished imetit-induced circulatory and oxygen uptake responses. Clobenpropit (4 micromol/kg i.a.) alone failed to affect the MBF, LDBF, AP, C and VO(2) values. Central administration of imetit (0.1 micromol i.c.v.) markedly increased AP and decreased MBF, LDBF, C and VO(2). Pretreatment with histamine H(3) receptor antagonist clobenpropit (0,4 micromol i.c.v.) diminished circulatory and metabolic responses to centrally injected imetit. Central histamine H(3) receptors blockade by clobenpropit evoked no significant changes in the mesenteric arterial and mucosal microcirculatory blood flow, intestinal metabolism and mean arterial pressure. We conclude that, peripheral histamine H(3) receptors when stimulated decreases vasoconstrictory tone of the mesenteric artery and precapillary structures and evokes increase of intestinal oxygen uptake. This might be in part due to inhibition of sympathetic postganglionic fibers vasopressor activity. Central histamine H(3) receptor stimulation activates vasoconstrictory sympathetic adrenergic system with possible involvement of other, presumably non-histaminergic receptors system. At basal conditions neither central nor peripheral histamine H(3) receptors are involved in the control of mesenteric macro- and microcirculation and intestinal oxygen consumption.

Effects of Ginkgo biloba on alertness and chemosensory function in healthy adults.

Ginkgo biloba is reported to enhance cognitive function in patients with selected neural disorders. Its effects in healthy, young adults are less well characterized. This work explored whether Ginkgo biloba could ameliorate decrements in alertness post-prandially and/or enhance chemosensory function. Both are functions that could be influenced by enhanced cerebral blood flow and neuronal metabolism, reported properties of the compound. A double-blind placebo-controlled study was conducted with 19 males and 20 females with a mean age of 23.6 +/- 5.4 years and mean weight of 70.0 +/- 1.9 kg. Participants were supplemented for 13 weeks with either Ginkgo biloba (mean dose 184.5 mg/d (range 130-234 mg/d)) or placebo and administered various alertness, performance, affective state and chemosensory tests at weeks 1, 5, 9 and 13. Participants did experience the post-prandial affective state decrement (i.e. post-lunch dip), but not the performance decrement. Performance on the chemosensory tests improved over the 13-week study. However, Ginkgo biloba was ineffective at alleviating the symptoms of the post-lunch dip or at enhancing taste and smell function.

The role of ascorbic acid and xylitol in etomidate-induced adrenocortical suppression in humans.

Etomidate-induced suppression of cortisol biosynthesis is a result of a blockade of 11-beta-hydroxylation in the adrenal gland, mediated by the imidazol radical of etomidate. Since the generation of steroids requires reductive and energy rich equivalents, the present study examined whether supplementation with ascorbic acid or xylitol, a major source of NADPH, could attenuate adrenal suppression by etomidate in human subjects by promoting the turnover rate of 11-beta-hydroxylase. During continuous etomidate/alfentanil anaesthesia for pelviscopic surgery 30 female patients received either Ringer's lactate, xylitol (0.25 g kg-1 h-1) or ascorbic acid (0.5 g h-1) intravenously (i.v.). The plasma concentrations of cortisol, aldosterone and dehydroepiandrosterone (DHEA) were recorded for 5 h after end of surgery and a stimulation with synthetic ACTH was performed. The results showed no evidence of a clinically relevant attenuating effect of ascorbic acid or xylitol on etomidate-induced adrenocortical suppression. However, the observed suppression of cortisol levels was not enough to allow an attenuating affect to be measured.