Study of hypoxia-induced expression of HIF-1α in retina pigment epithelium

It was shown that the expression of HIF-1α in retinal pigment epithelium increased under hypoxic conditions. Eight hours after the start of hypoxic exposure, the expression of HIF-1α reached the peak and sustained after 24-hour hypoxia. However, the morphology of PRE cells began to change and the expression of HIF-1α decreased after long-term (48-hour) hypoxia. Hypoxia-induced increase in the level of HIF-1α in RPE. It can be an important step in choriodal neovascularization. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 143, No. 3, pp. 293–297, March, 2007     

Moderate altitude but not additional endurance training increases markers of oxidative stress in exhaled breath condensate

Oxidative stress occurs at altitude, and physical exertion might enhance this stress. In the present study, we investigated the combined effects of exercise and moderate altitude on redox balance in ten endurance exercising biathletes, and five sedentary volunteers during a 6-week-stay at 2,800 m. As a marker for oxidative stress, hydrogen peroxide (H₂O₂) was analyzed by the biosensor measuring system Ecocheck™, and 8-iso prostaglandin F2α (8-iso PGF2α) was determined by enzyme immunoassay in exhaled breath condensate (EBC). To determine the whole blood antioxidative capacity, we measured reduced glutathione (GSH) enzymatically using Ellman’s reagent. Exercising athletes and sedentary volunteers showed increased levels of oxidative markers at moderate altitude, contrary to our expectations; there was no difference between both groups. Therefore, all subjects’ data were pooled to examine the oxidative stress response exclusively due to altitude exposure. H₂O₂ levels increased at altitude and remained elevated for 3 days after returning to sea level (p ≤ 0.05). On the other hand, 8-iso PGF2α levels showed a tendency to increase at altitude, but declined immediately after returning to sea level (p ≤ 0.001). Hypoxic exposure during the first day at altitude resulted in elevated GSH levels (p ≤ 0.05), that decreased during prolonged sojourn at altitude (p ≤ 0.001). In conclusion, a stay at moderate altitude for up to 6 weeks increases markers of oxidative stress in EBC independent of additional endurance training. Notably, this oxidative stress is still detectable 3 days upon return to sea level.

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The contribution of Max Gassmann and Katja Heinicke who are the senior authors was equivalent.     

Effects of HIF-1α on human gastric cancer cell apoptosis at different CO2 pressures

The effects and potential molecular mechanisms underlying carbon dioxide (CO₂) pneumoperitoneum on gastric cancer cell apoptosis are not fully understood. In this study, we assessed the effects of CO₂ pneumoperitoneum on the apoptosis of MKN-45 gastric cancer cells. Additionally, we investigated the role of HIF-1α in CO₂ pneumoperitoneum-induced apoptosis of gastric cancer cells. MKN-45 cells were cultured in CO₂ or air pneumoperitoneum at 0, 12 and 15 mmHg pressures for 4 h. We observed a change in cells morphology and increasing apoptotic ratios in MKN-45 cells when they were put into a 15 mmHg CO₂ pneumoperitoneum environment. However, there was no significant difference between the 0, 12 mmHg CO₂ pneumoperitoneum and the control groups. Exposure to 15 mmHg CO₂ pneumoperitoneum significantly enhanced the expression levels of HIF-1α and Bax, while it attenuated Bcl-2 expression levels. When we inhibited HIF-1α by small interfering RNA (siRNA), we found that the apoptotic ratio of MKN-45 cells decreased in 15 mmHg CO₂ pneumoperitoneum. This treatment markedly elevated Bcl-2 levels and decreased Bax expression. These data suggest that CO₂ pneumoperitoneum may accelerate the apoptosis of MKN-45 cells at higher pressures. HIF-1α is a crucial factor that affects gastric cancer cell apoptosis by downregulating the Bcl-2/Bax ratio.     

Thirteen days of “live high–train low” does not affect prooxidant/antioxidant balance in elite swimmers

We investigated the impact of 13 days of “living high–training low” (LHTL) on the antioxidant/prooxidant balance in elite endurance swimmers. Eighteen elite swimmers from the French Swimming Federation were submitted to a 13-day endurance training and divided into two groups: one group trained at 1,200 m and lived in hypoxia (2,500–3,000 m simulated altitude) and the second group trained and lived at 1,200 m. The subjects performed an acute hypoxic test (10 min at 4,800 m) before and 1 day after the training period. Plasma levels of advanced oxidation protein products (AOPP), malondialdehydes (MDA), ferric reducing antioxidant power (FRAP), and lipid-soluble antioxidants were measured before and after the 4,800 m tests. After the training, MDA and AOPP responses to the 4,800 m test were lower than before training for both groups (+10 vs. +2%; P = 0.01 for MDA and +80 vs. +14%; P = 0.01 for AOPP). Thirteen days of LHTL did not modify antioxidant status (FRAP and lipid-soluble antioxidants) despite intakes in vitamins A and E below the recommended daily allowances. The LHTL did not affect the antioxidant status in elite swimmers; however, the normoxic endurance training induced preconditioning mechanisms in response to the 4,800 m test.     

PPARα gene variation and physical performance in Russian athletes

Peroxisome proliferator-activated receptor α (PPARα) regulates genes responsible for skeletal and heart muscle fatty acid oxidation. Previous studies have shown that the PPARα intron 7 G/C polymorphism was associated with left ventricular growth in response to exercise. We speculated that GG homozygotes should be more prevalent within a group of endurance-oriented athletes, have normal fatty acid metabolism, and increased percentages of slow-twitch fibers. We have tested this hypothesis in the study of a mixed cohort of 786 Russian athletes in 13 different sporting disciplines prospectively stratified by performance (endurance-oriented athletes, power-oriented athletes and athletes with mixed endurance/power activity). PPARα intron 7 genotype and allele frequencies were compared to 1,242 controls. We found an increasing linear trend of C allele with increasing anaerobic component of physical performance (P=0.029). GG genotype frequencies in endurance-oriented and power-oriented athletes were 80.3 and 50.6%, respectively, and were significantly (P<0.0001) different compared to controls (70.0%). To examine the association between PPARα gene variant and fiber type composition, muscle biopsies from m. vastus lateralis were obtained and analyzed in 40 young men. GG homozygotes (n=25) had significantly (P=0.003) higher percentages of slow-twitch fibers (55.5±2.0 vs 38.5±2.3%) than CC homozygotes (n=4). In conclusion, PPARα intron 7 G/C polymorphism was associated with physical performance in Russian athletes, and this may be explained, in part, by the association between PPARα genotype and muscle fiber type composition.     

Interleukin-10 and Tumor Necrosis Factor-α Gene Polymorphisms in Tuberculosis

Tuberculosis (TB), caused by Mycobacterium tuberculosis, is an infectious disease in humans killing nearly three million people and eight million cases annually. The cytokines TNF-α and IL-10 have been implicated in the pathogenesis of TB. Certain single nucleotide polymorphisms within the promoter region of the IL10 and TNF genes have been associated with altered levels of circulating IL10 and TNF- α. We analyzed TNF-α (−308 G/A, −238 G/A, −376 G/A) and IL10 (−1,082 G/A, −819 C/T, −592 C/A) polymorphisms in 128 patients with TB and 80 healthy subjects using by amplification refractory mutation system-polymerase chain reaction (ARMS-PCR). A significant association was found between TB and −1,082 G allele (Pc: 0.000, O.R 2.22, 95% CI 1.45–3.41). Significant difference was observed in IL10 GCC and ACC haplotypes distribution between TB and control subjects (Pc: 0.000, O.R 2.22, 95% CI 1.45–3.41; Pc: 0.004, O.R 0.53, 95% CI 0.35–0.81). No statistically significant association was found between IL-10 −819 C/T, TNF-α 308 G/A, −238 G/A, −376 G/A polymorphisms, functional TNFα/IL-10 genotypes and TB. Our findings suggest that IL-10 108 2G/A alleles or haplotypes containing these alleles may influence the Th1/Th2 balance and hence may play a role in TB susceptibility and increase risk of developing disease. This polymorphism may be one of the many genetic factors affecting disease outcome.     

Differential association of SMC1α and SMC3 proteins with meiotic chromosomes in wild-type and SPO11-deficient male mice

SMC proteins are components of cohesin complexes that function in chromosome cohesion. We determined that SMC1α and SMC3 localized to wild-type mouse meiotic chromosomes, but with distinct differences in their patterns. Anti-SMC3 coincided with axial elements of the synaptonemal complex, while SMC1α was observed mainly in regions where homologues were synapsed. This pattern was especially visible in pachytene sex vesicles where SMC1α localized only weakly to the asynapsed regions. At diplotene, SMC3, but not SMC1α, remained bound along axial elements of desynapsed chromosomes. SMC1α and SMC3 were also found to localize along meiotic chromosome cores of Spo11 null spermatocytes, in which double-strand break formation required for DNA recombination and homologous pairing were disrupted. In Spo11−/− cells, SMC1α localization differed from SMC3 again, confirming that SMC1α is mainly associated with homologous or non-homologous synapsed regions, whereas SMC3 localized throughout the chromosomes. Our results suggest that the two cohesin proteins may not always be associated in a dimer and may function as separate complexes in mammalian meiosis, with SMC1α playing a more specific role in synapsis. In addition, our results indicate that cohesin cores can form independently of double-strand break formation and homologous pairing. This revised version was published online in July 2006 with corrections to the Cover Date.     

Reduced performance of male and female athletes at 580 m altitude

This study examined the effect of mild hypobaria (MH) on the peak oxygen consumption (O_2peak) and performance of ten trained male athletes [ (SEM); O_2peak = 72.4 (2.2) ml · kg⁻¹ · min⁻¹] and ten trained female athletes [O_2peak = 60.8 (2.1) ml · kg⁻¹ · min⁻¹]. Subjects performed 5-min maximal work tests on a cycle ergometer within a hypobaric chamber at both normobaria (N, 99.33 kPa) and at MH (92.66 kPa), using a counter-balanced design. MH was equivalent to 580 m altitude. O_2peak at MH decreased significantly compared with N in both men [− 5.9 (0.9)%] and women [− 3.7 (1.0)%]. Performance (total kJ) at MH was also reduced significantly in men [− 3.6 (0.8)%] and women [− 3.8 (1.2)%]. Arterial oxyhaemoglobin saturation (S_aO₂) at O_2peak was significantly lower at MH compared with N in both men [90.1 (0.6)% versus 92.0 (0.6)%] and women [89.7 (3.1)% versus 92.1 (3.0)%]. While S_aO₂ at O_2peak was not different between men and women, it was concluded that relative, rather than absolute, O_2peak may be a more appropriate predictor of exercise-induced hypoxaemia. For men and women, it was calculated that 67–76% of the decrease in O_2peak could be accounted for by a decrease in O₂ delivery, which indicates that reduced O₂ tension at mild altitude (580 m) leads to impairment of exercise performance in a maximal work bout lasting ≈ 5 min. Accepted: 30 July 1996     

Inhibition of histone deacetylase down-regulates the expression of hypoxia-induced vascular endothelial growth factor by rheumatoid synovial fibroblasts

Objective: To investigate the effect of FK228 on the in vitro expression of hypoxia-inducible factor-1 alpha (HIF-1α) and vascular endothelial growth factor (VEGF) by rheumatoid arthritis synovial fibroblasts (RASFs), and on the in vivo expression of VEGF and angiogenesis in the synovial tissue of mice with collagen-antibody-induced arthritis (CAIA). Methods: RASFs were stimulated with IL-1β and TNFα and then incubated under hypoxia (1 % O₂) with various concentrations of FK228. The effects of FK228 on the expression of HIF-1α and VEGF mRNA were examined by quantitative real-time PCR. Changes in HIF-1α protein expression and the secretion of VEGF protein into the culture medium were examined by Western blot analysis and ELISA, respectively. Immunohistochemical analysis was carried out to investigate the expression and distribution of VEGF in synovial tissues of CAIA mice. Results: The cytokine-stimulated expression of HIF-1α and VEGF mRNA was inhibited by FK228 in a dose-dependent manner. FK228 also reduced the expression of HIF-1α and VEGF protein. Intravenous administration of FK228 (2.5 mg/kg) suppressed VEGF expression, and also blocked angiogenesis in the synovial tissue of CAIA. Conclusion: FK228 may exhibit a therapeutic effect on RA by inhibition of angiogenesis through down-regulation of angiogenesis related factors, HIF-1α and VEGF. Received 28 February 2007; returned for revision 19 March 2007; accepted by J. Di Battista 11 July 2007     

The Transport Profile of Cytokines in Epidermal Equivalents Subjected to Mechanical Loading

Pressure ulcer risk assessment might be optimized by incorporating the soft tissue reaction to mechanical loading in the currently used risk assessment scales. Cytokines, like IL-1α, IL-1RA, IL-8, and TNF-α, might be used to determine this tissue reaction, since they are released after 24 h of mechanical loading of epidermal equivalents. In the current study, the release and transport of these cytokines with time was evaluated. Epidermal equivalents were subjected to 20 kPa for different time periods (1, 2, 4, 6, 8, 16, and 24 h). Compared to the unloaded control group, a significant increase was found for IL-1α (4.7-fold), IL-1RA (4.8-fold), and IL-8 (3.6-fold) release after 1 h loading. For TNF-α, the release was significantly increased after 4 h of loading (5.1-fold compared to the unloaded situation), coinciding with the first signs of gross structural tissue damage. These cytokine values were determined in the surrounding medium and a transport model was developed to evaluate the distribution of cytokines inside the culture. These simulations revealed that all IL-8 and TNF-α was released from the keratinocytes, whereas most of the IL-1α and IL-1RA remained inside the keratinocytes during the 24 h loading period. In conclusion, IL-1α, IL-1RA, and IL-8 appear promising biochemical markers for pressure ulcer risk assessment, since their release is increased after 1 h of epidermal loading and before the onset of structural tissue damage.     

Living high-training low: tolerance and acclimatization in elite endurance athletes

The “living high-training low” (LHTL) model is frequently used to enhance aerobic performance. However, the clinical tolerance and acclimatization process to this intermittent exposure needs to be examined. Forty one athletes from three federations (cross-country skiers, n=11; swimmers, n=18; runners, n=12) separately performed a 13 to 18-day training at the altitude of 1,200 m, by sleeping either at 1,200 m (CON) or in hypoxic rooms (HYP), with an O₂ fraction corresponding to 2,500 m (5 nights for swimmers and 6 for skiers and runners), 3,000 m (6 nights for skiers, 8 for swimmers and 12 for runners) and 3,500 m (6 nights for skiers). Measurements performed before, 1 or 15 days after training were ventilatory response (HVRe) and desaturation (ΔSaO₂e) during hypoxic exercise, an evaluation of cardiac function by echocardiography, and leukocyte count. Lake Louise AMS score and arterial O₂ saturation during sleep were measured daily for HYP. Subjects did not develop symptoms of AMS. Mean nocturnal SaO₂ decreased with altitude down to 90% at 3,500 m and increased with acclimatization (except at 3,500 m). Leukocyte count was not affected except at 3,500 m. The heart function was not affected by LHTL. Signs of ventilatory acclimatization were present immediately after training (increased HVRe and decreased ΔSaO₂e) and had disappeared 15 days later. In conclusion, LHTL was well tolerated and compatible with aerobic training. Comparison of the three patterns of training suggests that a LHTL session should not exceed 3,000 m, for at least 18 days, with a minimum of 12 h day⁻¹ of exposure.     

Effects of short-term corticoid ingestion on food intake and adipokines in healthy recreationally trained men

In order to test the hypothesis that short-term corticoid intake alters food intake, body composition and adipokines secretion in healthy volunteers with regular sport practice, nutrient intake was assessed in eight male athletes with and without prednisolone (PRED, 60 mg/day for 1 week) ingestion in a random, double blind, crossover design. Body weight, body composition, adipokines (i.e., leptin, adiponectin and TNF-α), insulin and blood glucose were determined before and at the end of each treatment. PRED did not induce any significant change in body weight, body composition or food intake. Insulin and TNF-α were not significantly altered with PRED compared to placebo but blood glucose, leptin and adiponectin concentrations at rest appear significantly increased after PRED treatment (P < 0.05). Our data show that 1 week glucocorticoid treatment does not promote obesity in recreationally trained men but further studies are necessary to understand its effects on the metabolically active hormones, leptin and adiponectin.     

Strength training improves cycling efficiency in master endurance athletes

The purpose of this study was to test the effect of a 3-week strength training program of knee extensor muscles on cycling delta efficiency in master endurance athletes. Nine master (age 51.5 ± 5.5 years) and 8 young (age 25.6 ± 5.9 years) endurance athletes with similar training levels participated in this study. During three consecutive weeks, all the subjects were engaged in a strength training program of the knee extensor muscles. Every week, they performed three training sessions consist of 10 × 10 knee extensions at 70% of maximal repetition with 3 min rest between in a leg extension apparatus. Maximal voluntary contraction torque (MVC torque) and force endurance (End) were assessed before, after every completed week of training, and after the program. Delta efficiency (DE) in cycling was evaluated before and after the training period. Before the training period, MVC torque, End, and DE in cycling were significantly lower in masters than in young. The strength training induced a significant improvement in MVC torque in all the subjects, more pronounced in masters (+17.8% in masters vs. +5.9% in young, P < 0.05). DE in cycling also significantly increased after training in masters, whereas it was only a trend in young. A significant correlation (r = 0.79, P < 0.01) was observed between MVC torque and DE in cycling in masters. The addition of a strength training program for the knee extensor muscles to endurance-only training induced a significant improvement in strength and cycling efficiency in master athletes. This enhancement in muscle performance alleviated all the age-related differences in strength and efficiency.     

Living high–training low: effect on erythropoiesis and aerobic performance in highly-trained swimmers

The “living high–training low” model (LHTL), i.e., training in normoxia but sleeping/living in hypoxia, is designed to improve the athletes performance. However, LHTL efficacy still remains controversial and also little is known about the duration of its potential benefit. This study tested whether LHTL enhances aerobic performance in athletes, and if any positive effect may last for up to 2 weeks after LHTL intervention. Eighteen swimmers trained for 13 days at 1,200 m while sleeping/living at 1,200 m in ambient air (control, n=9) or in hypoxic rooms (LHTL, n=9, 5 days at simulated altitude of 2,500 m followed by 8 days at simulated altitude of 3,000 m, 16 h day⁻¹). Measures were done before 1–2 days (POST-1) and 2 weeks after intervention (POST-15). Aerobic performance was assessed from two swimming trials, exploring and endurance performance (2,000-m time trial), respectively. Reticulocyte, serum EPO and soluble transferrin receptor responses were not altered by LHTL, whereas reticulocytes decreased in controls. In POST-1 (vs. before): red blood cell volume increased in LHTL only (+8.5%, P=0.03), tended to increase more in LHTL (+8.1%, P=0.09) than in controls (+2.5%, P=0.21) without any difference between groups (P=0.42) and 2,000-m performance was unchanged with LHTL. In POST-15, both performance and hematological parameters were similar to initial levels. Our results indicate that LHTL may stimulate red cell production, without any concurrent amelioration of aerobic performance. The absence of any prolonged benefit after LHTL suggests that this LHTL model cannot be recommended for long-term purposes.     

Heat Stress Protects Against Lung Injury in the Neutropenic,Endotoxemic Rat

The objective of this study is to determine if heat stress prior to endotoxemia diminishes cardiopulmonary dysfunction by attenuating the cytokine inflammatory response. Rats were assigned to either: 1) neutropenia; 2) heat; 3) neutropenia, LPS; or 4) heat, neutropenia, LPS. Heart rate, blood gases, and blood, lung lavage, and lung mRNA for tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and macrophage inflammatory protein (MIP)-2 were measured. Heat given before LPS resulted in a similar A-a O₂ gradient as the heat-alone and neutropenic groups (8 ± 8 versus 8 ± 7 versus 4 ± 3 mm Hg) and a lower A-a O₂ gradient when compared to the neutropenic, LPS rats (8 ± 8 versus 22 ± 8 mm Hg, p < 0.003). Blood, lung lavage, and lung mRNA for TNF-α, IL-1β, and MIP-2 were similar in the LPS rats regardless of heat. Heart rate was similar in both LPS groups but higher than non-LPS groups. Heat pretreatment attenuates lung injury in the neutropenic, endotoxemic rat but not by decreasing TNF-α, IL-1β, or MIP-2 in the lung. Heat prior to LPS did not prevent cardiac dysfunction in neutropenic rats.     

Variable severity of β-thalassemia patients of Eastern India: effect of α-thalassemia and XmnI polymorphism

Sixty-four thalassemia and E-β thalassemia patients were studied for factors that modulate the severity of the disease; i. e., mutation of β-globin gene, presence of α-deletion, and presence of an XnmI site at the −158 position of the Gγ gene. Presence of α-deletion and/or homozygosity for the XmnI site was in general associated with less-severe disease. About 12% of the patients harbored single α-gene deletion, and the gene frequency of the XnmI polymorphism in these patients is 0.48. Received: 16 January 2001 / Accepted: 18 September 2001