Effects of scrotal hyperthermia on Leydig cells in long-term: a histological,immunohistochemical and ultrastructural study in rats

We have previously demonstrated that scrotal hyperthermia induce Leydig cell (LC) damage in short-term. The objectives of this pilot study were to investigate morphological changes and regulation of steroidogenesis on LC in long-term and the time of observation were extended to investigate whether the LC would eventually make a recovery after scrotal hyperthermia. The rats were randomly allotted into one of four groups: A (control), B (70 days after scrotal hyperthermia), C (105 days after scrotal hyperthermia), D (140 days after scrotal hyperthermia); each group contain seven animals. Scrotal hyperthermia was carried out in a thermostatically controlled water bath at 43°C for 30 min once daily for six consecutive days. Control rats were treated in the same way, except the testes were immersed in a water bath maintained at 22°C. Hyperthermia applied rats were sacrificed under 50 mg/kg ketamine anaesthesia after 70, 105 and 140 days, and biopsy materials of testes were obtained for light and electron microscopic examinations. Morphologically normal and the number of testosterone positive LC was significantly higher in 140 days after last heat than all other heat treatment groups. In heat treated groups, a dilated smooth endoplasmic reticulum, swollen mitochondria, and vanished mitochondrial cristae were observed. In the 140 days after scrotal hyperthermia, the severities of degenerative changes of LC were less than that observed in the other heat treated groups. We conclude that, scrotal hyperthermia cause morphological damaging and impaired steroidogenesis in LC and recovery of these findings were noted first time in 140 days after the last heat treatment.     

The magnitude of physical exercise-induced hyperthermia is associated with changes in the intestinal permeability and expression of tight junction genes in rats

We investigated whether the magnitude of exercise-induced hyperthermia influences intestinal permeability and tight junction gene expression. Twenty-nine male Wistar rats were divided into four groups: rest at 24 °C and exercise at 13 °C, 24 °C or 31 °C. The exercise consisted of a 90-min treadmill run at 15 m/min, and different ambient temperatures were used to produce distinct levels of exercise-induced hyperthermia. Before the experimental trials, the rats were treated by gavage with diethylenetriaminepentaacetic acid labeled with technetium-99 metastable as a radioactive probe. The rats' core body temperature (T_CORE) was measured by telemetry. Immediately after the trials, the rats were euthanized, and the intestinal permeability was assessed by measuring the radioactivity of blood samples. The mRNA levels of occludin and zonula occludens-1 (ZO-1) genes were determined in duodenum samples. Exercise at 24 °C increased T_CORE to values close to 39 °C, without changing permeability compared with the resting trial at the same environment. Meanwhile, rats’ T_CORE exceeded 40 °C during exercise at 31 °C, leading to greater permeability relative to those observed after exercise in the other ambient temperatures (e.g., 0.0037%/g at 31 °C vs. 0.0005%/g at 13 °C; data expressed as medians; p < 0.05). Likewise, the rats exercised at 31 °C exhibited higher mRNA levels of ZO-1 and occludin genes than the rats exercised at 24 °C or 13 °C. The changes in permeability and gene expression were positively and significantly associated with the magnitude of hyperthermia. We conclude that marked hyperthermia caused by exercise in the warmer environment increases intestinal permeability and mRNA levels of tight junction genes.     

Heat shock increases levels of reactive oxygen species,autophagy and apoptosis

Hyperthermia is a promising anticancer treatment used in combination with radiotherapy and chemotherapy. Temperatures above 41.5 °C are cytotoxic and hyperthermia treatments can target a localized area of the body that has been invaded by a tumor. However, non-lethal temperatures (39–41 °C) can increase cellular defenses, such as heat shock proteins. This adaptive survival response, thermotolerance, can protect cells against subsequent cytotoxic stress such as anticancer treatments and heat shock (>41.5 °C). Autophagy is another survival process that is activated by stress. This study aims to determine whether autophagy can be activated by heat shock at 42 °C, and if this response is mediated by reactive oxygen species (ROS). Autophagy was increased during shorter heating times (<60 min) at 42 °C in cells. Levels of acidic vesicular organelles (AVO) and autophagy proteins Beclin-1, LC3-II/LC-3I, Atg7 and Atg12-Atg5 were increased. Heat shock at 42 °C increased levels of ROS. Increased levels of LC3 and AVOs at 42 °C were inhibited by antioxidants. Therefore, increased autophagy during heat shock at 42 °C (<60 min) was mediated by ROS. Conversely, heat shock at 42 °C for longer times (1?3 h) caused apoptosis and activation of caspases in the mitochondrial, death receptor and endoplasmic reticulum (ER) pathways. Thermotolerant cells, which were developed at 40 °C, were resistant to activation of apoptosis at 42 °C. Autophagy inhibitors 3-methyladenine and bafilomycin sensitized cells to activation of apoptosis by heat shock (42 °C). Improved understanding of autophagy in cellular responses to heat shock could be useful for optimizing the efficacy of hyperthermia in the clinic.     

Anticancer effects of 6-<Emphasis Type="Italic">o</Emphasis>-palmitoyl-ascorbate combined with a capacitive-resistive electric transfer hyperthermic apparatus as compared with ascorbate in relation to ascorbyl radical generation

The aim of the present study is to determine the anti-proliferative activity of 6-o-palmitoyl-l-ascorbic acid (Asc6Palm) that is a lipophilic derivative of l-ascorbic acid (Asc), on human tongue squamous carcinoma HSC-4 cells by combined use of hyperthermia in comparison to Asc. Asc6Palm or Asc were administered to HSC-4 cells for 1 h, to which hyperthermia at 42 °C was applied for initial 15 min. After further 1–72 h incubation at 37 °C, cell proliferation was determined with Crystal Violet staining. Ascorbyl radical (AscR) in HSC-4 cell suspension was measured by electron spin resonance (ESR), and cell morphology was observed with scanning electron microscopy (SEM). At 37 °C, 4 mM Asc or 0.35 mM Asc6Palm were enough to suppress proliferation of HSC-4 cells. By combined use of hyperthermia at 42 °C, cell proliferation was decreased when compared to 37 °C. After Asc of 4 mM was incubated with HSC-4 cell suspensions at 37 °C or 42 °C for 0–180 min, the signal intensity of ascorbyl radical (AscR) by ESR was not different regardless of the presence or absence of cells at 37 °C, whereas AscR signal was enlarged in the presence of HSC-4 cells at 42 °C. It was suggested that oxidation of Asc occurred rapidly in HSC-4 cells by hyperthermia, and thereby enhanced the anti-proliferative activity. By SEM observation, the surface of HSC-4 cells treated with Asc6Palm revealed distinct morphological changes. Thus, the combined regimen of Asc6Palm and hyperthermia is expected to exert a marked antitumor activity.     

The impact of oral ciprofloxacin on the structure and functions of rat gastric mucosa

Ciprofloxacin (CPX), is a fluoroquinolone antibiotic used to treat a number of gram-negative and gram-positive bacterial infections. Ciprofloxacin can cause severe side effects, ranging from tendon problems, nerve damage, to serious mood or behavior changes.The purpose of this study was to investigate how ciprofloxacin affects gastric cell lines in rats with a distinctive emphasis on physiological, histopathological, and bacteriological changes. Male albino rats (n = 21) were distributed into three groups; control, CPX, and CPX-withdrawal groups. The treated rats were given CPX tablets (12.5 mg/kg) dissolved in carboxymethyl cellulose (CMC) 0.5% orally once daily via gavage for sixty consecutive days. Control rats received only the vehicle. The withdrawal group was treated for 60 days and the drug was withdrawn for another sixty days. After completion of the experiment, all rats were sacrificed and gastric tissues were treated for light, immunohistochemical, and scanning electron microscopic examination. Image J software was used to measure immune-labeled gastric epithelial cells. Blood samples were also collected for H. Pylori immunoglobulins IgM, IgA, and IgG. Results showed that treated rats acquired significantly strongly positive tumor necrosis factor (TNFα) and significant reduction of serum level of H. pylori IgM, IgA, and IgG in all the study groups. It could be concluded that prolonged oral CPX administration to albino rats changes the gastric mucosal architecture and bacteriology.     

Effects of acute and chronic testicular hyperthermia on levels of testosterone and corticosteroids in plasma of boars

Heat is known to depress spermatogenesis in the boar, but there is little quantitative evidence on its effects on testicular steroidogenesis in this species. The studies reported here examine the effects of short-term and chronic testicular hyperthermia on levels of testosterone (T) and corticosteroids (C) in plasma of Large-White (LW) boars.In examining effects of acute heating, three mature LW boars were maintained at 23°, 35° and 23°C ambient during three consecutive 24-h periods. Blood samples were collected hourly and levels of T and C in plasma determined. Prior to heating, plasma T levels varied diurnally (P<0.05) about a 24-h mean value of 2.78 nM. During heating at 35°C, and recovery at 23°C, mean plasma T levels remained unchanged (P>0.05) but there was a loss of diurnal rhythm. Mean 24-h plasma C levels did not change during heating (20.8 nMat 23°C, 20.2 nMat 35°; P>0.05), but fell (P<0.05) to 8.3 nM during the recovery period at 23°C.Effects of chronic heating on testis function were investigated by determining T and C concentrations in peripheral plasma of unilateral cryptorchid boars in which the scrotal testis was removed shortly after birth. Blood samples were drawn hourly, for 24 h, from each animal at about 10 months of age. The boars were then treated, i.v., with 700 IU hCG and blood samples collected frequently for 12 h. Mean plasma T levels before and after hCG treatment were 1.94 and 3.71 nM respectively, the difference between these levels being significant (P<0.05). At the same time, comparison was made with four normal littermates, hemicastrated at 3 days of age and heated to maintain testis temperature near 38°C. Mean plasma T levels in these boars increased (P<0.05) from 5.90 nM before, to 26.5 nM after hCG treatment, both levels being higher (P<0.05) than corresponding values for the hemicastrate cryptorchid animals. Levels of C in plasma increased (P<0.05) in the heated-scrotal boars following hCG treatment but decreased (P<0.05) in the cryptorchid animals. Histological comparison of testicular tissue from the scrotal and cryptorchid animals in this experiment revealed hypertrophy of Leydig cells in the abdominal testes.It is concluded that acute testicular hyperthermia (to c. 38°C) does not result in significant depression in mean plasma T levels of boars. However, chronic heating of testes at 38°C is associated with lower basal levels of T in peripheral plasma and an impaired response of plasma T concentrations following gonadotrophic stimulation.     

Morphometrical and intracellular changes in rat ovaries following chronic administration of ghrelin

The aim of our investigation was to examine the influence of chronic administration of ghrelin on the rat ovarian state. Morphometrical and intracellular changes in the ovary of 35-d female Wistar rats after sc injection of 1 nmol of ghrelin for 10 consecutive days were studied. Control animals (n = 10) were injected with normal saline using similar method. The ovaries were collected on days 1 and 6 after last injection from each group and subjected to light microscopic morphometric and electron microscopic analysis. It was demonstrated that the number of corpora lutea was significantly lower and the number of ovarian follicles was higher in the treated group on days 1 and 6, than in control (P < 0.01). Moreover, the mean diameter of each follicle, corpora lutea, luteal cell, theca layer, oocyte and zona plucida, but not of granulosa layer, as well as the whole ovarian volume were significantly lower in the treated animals at days 1 and 6 (P < 0.05). Electron microscopic analysis also indicated some intracellular changes associated with apoptosis and cell death such as presence of secondary lysosome, apoptotic bodies, nuclear chromatin condensation as well as margination, nuclear segmentation and vacuolization of cytoplasm of granulosa and theca cells. Our observations provides novel evidences for inhibitory influence of ghrelin on rat ovarian structures and, therefore, for the role of ghrelin as suppressor of female reproductive system.     

The induction of microphthalmia, encephalocele, and other head defects following hyperthermia during the gastrulation process in the rat

The aim of this study was to ascertain whether there is a period during early embryonic development of the rat that is particularly sensitive to hyperthermia. Pregnant Sprague-Dawley rats were partially immersed in a water bath at 43.5 degrees C until their core temperatures, monitored by a rectal thermistor probe, were elevated to 43.5 degrees C. The procedure was repeated 6 hours later. The regimen of two heatings was performed over a range of development from early gastrulation (8 days 18 hours) to about the 12 somite stage (10 days 18 hours). The rats were killed on days 17-19 and the fetuses were examined. Each group contained a minimum of five litters. The main teratogenic effect of the hyperthermia was the induction of one or more head defects, notably microphthalmia, encephalocele (either a single, large, parietal encephalocele or multiple small protuberances), and maxillary hypoplasia. Microphthalmia was the most common defect with approximately 90% of surviving fetuses having small eyes when heating occurred between 9 days 6 hours and 10 days 0 hours (9.06 and 10.00). Encephaloceles were induced by heating between 9.00 and 10.00 with a peak sensitivity between 9.12 and 9.18 when 57% of surviving fetuses were affected. Maxillary hypoplasia resulted from heating between 9.06 and 10.06 with up to 20% of surviving fetuses being affected. Control rats were exposed to the same experimental procedure in a water bath at 38 degrees C on 9.12 and 9.18, the gestational time most sensitive to hyperthermia induced malformations. There were no abnormal fetuses in the controls. The critical period identified spans 9 days 6 hours to 10 days 0 hours gestational age. In developmental terms this includes a large proportion of the gastrulation process.     

Effect of exposure to high temperatures in the excretion of cadmium and lead

ObjectiveThis study aims to observe the effect on urine and sweat excretion levels of cadmium (Cd) and lead (Pb) in healthy men in a maximum incremental test until exhaustion and repeated exposure to heat.Methodstwenty-nine adult men divided into control group (CG; n = 14) and experimental group (EG; n = 15) performing two maximum tests until exhaustion in normothermia (22 °C) and hyperthermia (42 °C). EG experienced 9 sessions of heat exposure at high temperatures (100 °C) (HEHT). After the nine sessions, the initial tests were repeated in both groups. Urine samples were collected before and after each test. After the hyperthermia tests, sweat samples were gathered.ResultsUrinary Cd increased after initial tests in GC and in hyperthermia in EG (p < 0.05). Urinary excretion of Pb rose after HEHT (p < 0.05). Pb in sweat was higher in EG than in CG after HEHT (p < 0.05).ConclusionHeat exercise and constant exposure to heat can be a valid method to increase the excretion of toxic metals.     

Cholestasis induced nephrotoxicity: The role of endogenous opioids

AimsElevated levels of endogenous opioids play a pivotal role in several deleterious consequences of cholestasis. Renal dysfunction occurs in cholestasis but its exact mechanism is still unknown. In this study, we investigated the role of endogenous opioids in cholestasis induced nephrotoxicity.Main methodsThirty-five rats were divided into five groups. In groups 1 and 2 BDL rats received either daily subcutaneous 20 mg/kg of naltrexone or its vehicle, for 7 days after BDL. In groups 3 and 4, BDL or Sham rats received no injections. In group 5, normal rats received subcutaneous injections of 20 mg/kg/day of naltrexone for 7 days. At the 7th day, 24 h urine was collected to measure urinary N-acetyl-β-D-glucosaminidase (NAG) as an early marker of renal tubular injury. Kidney samples were then collected for light and electron microscopic studies.Key findingsBDL significantly increased NAG activity compared to sham groups. Naltrexone significantly reversed NAG activity to normal levels in BDL animals. Naltrexone treatment in BDL animals also significantly reversed ALT and AST to their normal levels. In light and electron microscopic studies, there were significant structural alterations in BDL samples, which were mostly prevented in naltrexone treated BDL animals.SignificanceSignificant changes in urinary NAG activity and renal morphology of cholestatic rats were reversed by naltrexone treatment. These results suggest a possible role for endogenous opioids in inducing cholestatic nephrotoxicity.     

Effects of Temperature and Culture Media on Vegetative Growth of an Entomopathogenic Fungus <Emphasis Type="Italic">Isaria</Emphasis> sp. (Hypocreales: Clavicipitaceae) Naturally Affecting the Whitefly, <Emphasis Type="Italic">Bemisia tabaci</Emphasis> in Texas

The effects of temperature and mycological media on mycelial growth and estimates of spore production of an indigenous entomopathogenic fungus, Isaria sp., found during natural epizootics on whiteflies in the Lower Rio Grande Valley of Texas, were investigated. The radial growth (mm/day) of Isaria sp. as a function of temperature fits a linear model; with faster growth on Sabouraud dextrose agar with yeast extract, SDAY slopes (0.23) than on Sabouraud maltose agar, SMA slopes (0.14) from 20 to 30°C, with an optimal temperature of 30°C (SDAY: 4.1 mm, SMA: 3.1 mm). Moderate growth occurred at 25°C (SDAY: 3.4 mm, SMA: 2.7 mm). Growth was lowest at 20°C (SDAY: 1.9 mm, SMA: 1.8 mm). No fungal growth was observed at 35°C and 40°C. However, when Isaria sp. was exposed to 35°C for the first 7 days, it could recover and grow when transferred to 25°C (SDAY: 3.5 mm, SMA: 2.8 mm). No recovery or growth occurred after transfer from 40°C to 25°C. The average conidial production on SDAY after 20 days incubation at 25°C and a photoperiod of 14:10 h light: dark was 1.2 × 10⁸ conidia/cm² with 100% spore viability. When compared on SDAY at 25°C, the radial growth rate of I. javanica ex type CBS 134.22 (5.1 mm/day) was greater than seven Isaria isolates including Isaria sp.; but maximum growth rates were similar among all related Isaria isolates (90–97%). The Isaria sp. fungus tolerates high temperatures (35°C), suggesting that it is naturally selected for the subtropical semi-arid environment, where it could serve as an important natural control agent of the sweet potato whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) biotype B, one of the most invasive and economically damaging insects to agriculture. Mention of trade names or commercial products in this publication is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U. S. Department of Agriculture.     

Influence of a high-temperature programme on serum,urinary and sweat levels of selenium and zinc

IntroductionThe effect of hyperthermia on the antioxidant system in the human organism is well known.AimThe objective of this study was to observe the effects of heat on the concentration of Se and Zn, elements related to antioxidant systems.MethodsTwenty-nine subjects voluntarily participated in this study. They were divided into a control group (CG; n = 14) and an experimental group (EG; n = 15). All of them underwent two incremental tests until exhaustion in normothermia (22 °C, 20–40%RH) and hyperthermia (42 °C, 20–40%RH). EG experienced nine sessions of repeated heat exposure at high temperatures (100 °C, 20%RH) for three weeks (HEHT). After the intervention, the initial measurements were repeated. Urine and blood samples were collected before and after each test. Additionally, sweat samples were collected after tests in hyperthermia.ResultsThere were no significant changes in serum. An increase in the elimination of Zn and Se in EG was observed in urine after HEHT (p < .05). The elimination of Zn by sweating decreased after HEHT in EG (p < .05).ConclusionsExposure to heat at high temperatures increases the urinary excretion of Se and Zn.     

Effects of temperature change on the innate cellular and humoral immune responses of orange-spotted grouper Epinephelus coioides and its susceptibility to Vibrio alginolyticus

Orange-spotted grouper Epinephelus coioides held at 27 °C were then further cultured at 19, 27 (control), and 35 °C, and were examined for innate cellular and humoral responses after 3–96 h. The total leucocyte count, respiratory burst, and phagocytic activity significantly decreased 3, 48, and 96 h after fish were transferred to 19 and 35 °C. Both the alternative complement pathway (ACH₅₀) and the lysozyme activity significantly decreased at 3–96 h after fish were transferred to 19 and 35 °C. In another experiment, groupers reared at 27 °C at 34‰ salinity were injected with Vibrio alginolyticus grown in tryptic soy broth (TSB) at a dose of 2.3 × 10⁹ colony-forming units (cfu) fish^?1, and then further reared in water temperatures of 19, 27 (control), and 35 °C. The cumulative mortalities of V. alginolyticus-injected fish held in 19 and 35 °C were significantly higher than that of injected fish held in 27 °C. Resistance had decreased after 12 h for the challenged grouper held at 35 °C. All injected fish held in 19 °C had died after 72 h. It was concluded that at 12 h after transfer of grouper from 27 to 19 and 35 °C, immunity was suppressed and resistance against V. alginolyticus had decreased.     

Temperature and storage time strongly affect the germination success of perennial Euphorbia species in Mediterranean regions

This study aims to explore the effect of environmental factors (temperature, light, storage time) on germination response and dormancy patterns in eight Mediterranean native wildplants, belonging to the Euphorbia L. genus. In detail, we considered E. amygdaloides subsp. arbuscula, E. bivonae subsp. bivonae, E. ceratocarpa, E. characias, E. dendroides, E. melapetala, E. myrsinites, and E. rigida. We collected seeds from natural plant populations and performed germination assays in climatic chambers at seven constant temperatures (from 5 to 35°C, with 5°C increments), and four fluctuating temperature regimes (8/15, 8/20, 8/25, and 8/30°C, with a 12/12 hr thermoperiod). Germination assays were set up both in dark (D) and in light/dark conditions (L/D, 12/12 hr photoperiod), after short and long seed storage (SS around 30 days and LS around 150 days). For all these species, except E. amygdaloides subsp. arbuscula, results show that the final germinated proportions were improved by a long storage period (>150 days), which supports the existence of nondeep physiological dormancy. Optimal temperature levels ranged from 14.3 to 21.3°C and base temperatures ranged from 5.6 to 12.1°C, while ceiling temperatures from 25.6 to 34.7°C. For none of these species, germinations were favored by an alternating daily temperature regime, while in several instances, germinations were quicker and more complete in darkness, than in an alternating light/dark regime. In some instances, extreme temperature levels (5 and 30°C) induced dormancy and germinations did not resume when seeds were exposed at optimal temperature levels. Results are discussed in terms of the dynamics of emergences and how this might be affected by climate changes.     

Reproduction,survival, and life table parameters of the predatory mite <Emphasis Type="Italic">Cheyletus malaccensis</Emphasis> (Acari: Cheyletidae) at various constant temperatures

Reproduction, survival, and life table parameters of the predatory mite Cheyletus malaccensis Oudemans were evaluated at six constant temperatures: 17.5, 20, 25, 30, 32.5 and 35°C, feeding on Tyrophagus putrescentiae (Schrank). Preoviposition period of fertilized and virgin females varied with temperature from ca. 9 days at 17.5°C to ca. 1.5 day at 32.5°C and then increased to ca. 3 days at 35°C. Virgin female oviposition period was significantly shorter than for fertilized females at the temperatures examined with the exception of 17.5°C. The mean total number of eggs per fertilized (169.7 ± 6.6) and virgin female (60.7 ± 4.3) was highest at the temperature of 30°C. The data indicated a significant positive and nearly doubling effect of fertilization on female fecundity at the temperatures examined with the exception of 17.5°C. Age-specific fecundity was described by a temperature dependent model from which the maximum daily fecundity rate was estimated for fertilized and virgin females at 10.3 (at 30°C) and 6.8 (at 32.5°C) eggs/female, respectively. Virgin female longevity was significantly shorter than for fertilized females at 20, 30 and 32.5°C, and decreased from ca. 57 days at 17.5°C to ca. 17 days at 35°C. The Weibull function that was used to describe the age specific survival of fertilized and virgin females produced excellent fits to the survival data. Estimates of intrinsic rate of increase, net reproductive rate, mean generation time, doubling time and finite rate of increase, were obtained. The r_m value increased with temperature from 0.03 (day⁻¹) at 17.5°C to 0.21 (day⁻¹) at 32.5°C, after which it decreased to 0.15 (day⁻¹) at 35°C. These data indicate that C. malaccensis can reproduce at temperatures between 17.5 and 35°C and can be used for biological control of astigmatid mites within the temperature range where the pest occurs.     

Sweat rate and sweat composition during heat acclimation

The purpose of this study was to determine local sweat rate (LSR) and sweat composition during heat acclimation (HA). For ten consecutive days of HA, eight participants cycled in 33 °C and 65% relative humidity at an intensity such that a rectal temperature of 38.5 °C was reached within ~40 min, followed by a 60-min clamp of this rectal temperature (i.e., controlled hyperthermia). Four participants extended HA by a 28-day decay period and five consecutive days of heat re-acclimation (HRA) using controlled hyperthermia. Sweat from the upper arm and upper back was collected three times during each heat exposure session. LSR and sweat sodium, chloride, lactate, and potassium concentrations were determined. Relative to HA day 1, LSR was increased at the final day of HA (day 10) (arm: +58%, P < 0.001; back: +36%, P < 0.05). Concentrations of sodium, chloride, and lactate significantly (P < 0.05) decreased to ~60% at HA day 10 compared to day 1 on the arm and back. Potassium concentration did not significantly differ on HA day 10 compared to day 1 (arm: +11%, P > 0.05; back: +8%, P > 0.05). The induction patterns of the sudomotor adaptations were different. Whilst LSR increased from HA day 8 on the arm and from HA day 7 on the back, sodium and chloride conservation already occurred from HA day 3 on both skin sites. Lastly, the sweat lactate reduction occurred from HA day 6 on the arm and back. Initial evidence is provided that adaptations were partly conserved after decay (28 days) and that a 5-day HRA may be sufficient to restore HA adaptations. In conclusion, ten days of exercise-induced HA using controlled hyperthermia led to increases in LSR and concomitant reductions of sweat sodium, chloride, and lactate concentrations, whilst potassium concentrations remained relatively constant.     

Germination of Tagetes minuta L. I. Temperature Effects

Initial studies have indicated that Tagetes minuta achenes haveboth a temperature and a light requirement for germination.Temperatures tested were 10, 20, 25, 30 and 35 °C. Germinationwas optimal at 25 °C under white light conditions. Underthese conditions 100 per cent of achenes germinated within 7days of imbibition. There was no germination at 10 or 35 °Ceither in the light or in the dark. Achenes imbibed and incubatedat 35 °C for 4 days showed no visible signs of germinationbut on transfer to 25 °C, 100 per cent of these achenesgerminated within 24 h. Furthermore, achenes given this hightemperature (35 °C) treatment could be dried at 25 °C,re-imbibed at 25 °C and again 100 per cent of achenes germinatedwithin 24 h of re-imbibition. This rapid germination responsefollowing removal from the high temperature regime could alsobe induced by transfer to temperatures of 20 °C or 20 °C(16 h) alternating with 10 °C (8 h). Tagetes minuta L., weed seeds, germination, temperature, light     

Specific induction of a 72-kDa heat shock protein protects esophageal mucosa from reflux esophagitis

AimsThe aim of this study is to investigate the expression and cytoprotective function of a 72-kDa heat shock protein (HSP72) using a reflux esophagitis model in rats.Main methodsExpression of HSP60, HSP72, and HSP90 in rat esophageal mucosa was evaluated by Western blot analysis before and after hyperthermia (42.5 °C, 20 min). Rats received the operation to produce reflux esophagitis with or without pretreatment with hyperthermia to induce HSPs. The esophageal mucosal damage was evaluated 12 h after the operation.Key findingsExpression of HSP72 was significantly increased by hyperthermia in rat esophageal mucosa. Reflux esophagitis was dramatically prevented when HSP72 was preinduced by hyperthermia. Furthermore, activation of TNF-α and IL-1β in esophageal mucosa was also suppressed.SignificanceThese results suggested that hyperthermia protects the esophageal mucosa in reflux esophagitis model by inducing HSP72 and suppressing proinflammatory cytokine activation. These findings might suggest that HSP-inducing therapy could be a novel and unique therapy for reflux esophagitis.     

Effect of maturation conditions on light and temperature requirements during seed germination of Citrullus colocynthis from the Arabian Desert

• Seed germination of Citrullus colocynthis, as in many other species of Cucurbitaceae, is inhibited by light, particularly at low temperatures. Germination response to light and temperature has been attributed to day length and temperature during seed maturation. This study assessed the effects of these factors on the germination response of C. colocynthis to temperature and light quality.
• Ripe fruits were collected from natural habitats during December and February and germinated at three temperatures (15/25, 20/30 and 25/35 °C) in five light treatments (dark, white light and Red:Far Red (R:FR) ratios of 0.30, 0.87 and 1.19). Additionally, unripe fruits were also collected from natural habitats and completed their maturation in growth chambers under different day lengths (6, 16 and 24 h of darkness) at 10/20 °C, and in darkness at both 10/20 °C and 25/35 °C. Mature seeds of the different treatments were germinated in the same five light treatments at 15/25 °C.
• Germination was significantly higher in the dark than that in any light treatment. Seeds matured at higher temperatures (i.e. seeds from the December collection and those matured at 25/35 °C) had significantly higher germination than those matured at lower temperatures (i.e. seeds from the February collection and those matured at 10/20 °C). Dark germination was significantly higher for the December collection than for the February collection. Seeds of the two collections germinated in the dark only at 15/25 °C. However, seeds matured in a growth chamber at 10/20 °C in darkness germinated at 15/25 °C in all light treatments, except for the R:FR ratio 0.30. Seeds of the different treatments failed to germinate in FR‐rich light.
• This study demonstrates that both temperature and day length during seed maturation play significant roles in the germination response of C. colocynthis. Additionally, the dark requirement for germination is likely beneficial for species with the larger seeds, such as C. colocynthis, which produce bigger seedlings that are able to emerge from deep soils and are competitively superior under dense vegetation and resource‐limited conditions.

     

A simple cryopreservation protocol of <Emphasis Type="Italic">Dioscorea bulbifera</Emphasis> L. embryogenic calli by encapsulation-vitrification

Embryogenic calli of Dioscorea bulbifera L. were successfully cryopreserved using an encapsulation-vitrification method. Embryogenic calli were cooled at 6°C for 5 days on solid MS medium (Murashige and Skoog 1962) containing 2 mg L⁻¹ Kinetin (Kn), 0.5 mg L⁻¹ α-naphthalene acetic acid (NAA) and 0.5 mg L⁻¹ 2,4-dichlorophenoxy-acetic acid (2,4-D). These were prior precultured on liquid basal MS medium enriched with 0.75 M sucrose at 25 ± 1°C for 7 days. Embryogenic calli were osmoprotected with a mixture of 2 M glycerol and 1 M sucrose for 80 min at 25°C and dropped in a 0.1 M CaCl₂ solution containing 0.4 M sucrose at 25 ± 1°C. After 15 min of polymerization, Ca-alginate beads (about 4 mm in diameter) were dehydrated for 150 min at 0°C in a PVS₂ solution [30% glycerol, 15% ethylene glycol, and 15% dimethyl sulfoxide (w/v)] containing 0.5 M sucrose. The encapsulated embryogenic calli were then plunged directly into LN (liquid nitrogen) for 1 h. After rapid thawing in a water bath (37°C; 2 min), the beads were washed 3 times at 10-min intervals in liquid basal MS medium containing 1.2 M sucrose. Following thawing, the embryogenic calli were transferred to fresh solid basal MS media supplemented with Kn 2 mg L⁻¹, 0.09 M sucrose and 0.75% (w/v) agar (embryoid induction medium) and cultured under light conditions of 12-h photoperiod with a light intensity of 36 μmol m⁻² s⁻¹ provided by white cool fluorescent tubes after a 2-day dark period at 25 ± 1°C. After 30 days, the embryoids developed from embryogenic calli were transferred to fresh solid basal MS media supplemented with Kn 2 mg L⁻¹, NAA 0.5 mg L⁻¹, 3% (w/v) sucrose and 0.75% (w/v) agar (regeneration medium). After 60 days, the embryogenic calli developed normal shoots and roots. No morphological abnormalities were observed after plating on the regeneration medium. The survival rate of encapsulated vitrified embryogenic callus reached over 70%. This encapsulation-vitrification method appears promising as a routine and simple method for the cryopreservation of Dioscorea bulbifera embryogenic callus.