急性冷应激对牦牛乳腺上皮细胞 HSP70 mRNA 表达的影响

本文主要研究了急性冷应激对牦牛乳腺上皮细胞热休克蛋白７０ （Heat stress protein,ＨＳＰ７０） 表达量的影响。采用实时荧光定量ＰＣＲ 技术,以急性冷刺激１０℃ 为典型研究环境,分析了ＨＳＰ７０ ｍＲＮＡ 表达的变化规律。结果显示,乳腺上皮细胞在１０℃分别冷处理２ ｈ、４ ｈ、６ ｈ 和８ ｈ,其ＨＳＰ７０ ｍＲＮＡ 的表达量变化均不显著（Ｐ ＞０. ０５）;分别在１０℃冷处理２ ｈ、４ ｈ、６ ｈ 和８ ｈ,再复温培养４ ｈ,ＨＳＰ７０ ｍＲＮＡ 的表达量均极显著增加（Ｐ ＜０. ０１）,于６ ｈ 达到峰值;在１０℃先冷处理４ ｈ,然后分别复温２ ｈ、４ ｈ、６ ｈ 和８ ｈ,ＨＳＰ７０ ｍＲＮＡ 的表达量亦均显著增加（Ｐ ＜０. ０１）,并于４ ｈ 达到峰值。结论：急性冷应激诱导牦牛乳腺上皮细胞ＨＳＰ７０ 表达量的增加不是发生在冷处理过程中,而是发生在复温过程中,并且在一定范围内随冷处理时间的增长表达量增高。     

Seasonal variation in HSP70 expression and oxidative stress in skin of zebu (Tharparkar) and crossbred (Karan Fries) cattle under tropical climate

The study aimed to evaluate inducible HSP70 (HSP70.1 and HSP70.2) gene expression and oxidative stress status in skin of cattle during different seasons. Ten each of Tharparkar (zebu) and Karan Fries (crossbred) heifers were selected from NDRI herd, Karnal. Animals were maintained under standard managemental practices followed at the farm. Skin biopsies were aseptically collected from each animal during winter, spring, and summer. Real time PCR was performed to examine HSP70 expression. Reactive oxygen species (ROS) and antioxidant enzymes (SOD and CAT) were determined by ELISA. In both the breeds, significantly higher (p < 0.05) levels of HSP70 expression, ROS, caspases, and antioxidant enzymes were observed during summer followed by winter and spring. Breeds showed no significant difference during winter and spring. During summer, HSP70 expression, ROS, and antioxidant enzymes were higher (p < 0.05) in Karan Fries than Tharparkar, whereas caspases levels were higher in Tharparker than Karan Fries. The study concludes that levels of HSP70 expression, ROS, caspases, and antioxidant enzymes in skin of cattle were strongly affected by seasonal change in temperature. Differences exist in skin tissue thermotolerance of Tharparkar and Karan Fries cattle. This might be an efficient and centrally important mechanism for better adaptability of zebu cattle to heat stress.     

Restriction fragment length polymorphisms at the heat shock protein HSP70 gene(s) in pigs*

Pigs from a population consisting of eight US breeds or strains and three Chinese breeds were examined by restriction fragment length polymorphism (RFLP) analysis of the heat shock protein HSP70 gene(s). Limited polymorphisms with PstI and PvuII restriction enzymes were observed, but there were no polymorphisms with BomIII and BglI.     

Seasonal variation in expression pattern of genes under HSP70

Heat shock protein 70 (HSP70) is one of the most abundant and best characterized heat shock protein family that consists of highly conserved stress proteins, expressed in response to stress, and plays crucial roles in environmental stress tolerance and adaptation. The present study was conducted to identify major types of genes under the HSP70 family and to quantify their expression pattern in heat- and cold-adapted Indian goats (Capra hircus) with respect to different seasons. Five HSP70 gene homologues to HSPA8, HSPA6, HSPA1A, HSPA1L, and HSPA2 were identified by gene-specific primers. The cDNA sequences showed high similarity to other mammals, and proteins have an estimated molecular weight of around 70 kDa. The expression of HSP70 genes was observed during summer and winter. During summer, the higher expression of HSPA8, HSPA6, and HSPA1A was observed, whereas the expression levels of HSPA1L and HSPA2 were found to be lower. It was also observed that the expression of HSPA1A and HSPA8 was higher during winter in both heat- and cold-adapted goats but downregulates in case of other HSPs. Therefore, both heat and cold stress induced the overexpression of HSP70 genes. An interesting finding that emerged from the study is the higher expression of HSP70 genes in cold-adapted goats during summer and in heat-adapted goats during winter. Altogether, the results indicate that the expression pattern of HSP70 genes is species- and breed-specific, most likely due to variations in thermal tolerance and adaptation to different climatic conditions.     

Exogenous HSP70 becomes cell associated,but not internalized,by stressed arterial smooth muscle cells

Summary Cell death within atherosclerotic plaques leads to necrosis and rupture, resulting in vascular occlusion. We have previously demonstrated that addition of exogenous 70 kDa heat shock protein (HSP70) to arterial smooth muscle cells (aSMCs) in vitro can protect against toxins that may initiate necrosis. To determine whether exogenous HSP70 enters aSMCs or acts from outside cells to preserve viability, cultured rabbit aSMCs were stressed by serum deprivation and treated with fluorescently labeled (7-aminomethyl-4-coumarin-3-acetate) or¹²⁵I-radiolabeled HSP70. Cell-associated HSP70 was analyzed using Western blotting, fluoresence spectroscopy, and gamma counting/autoradiogarphy. Surface binding of HSP70 to aSMCs was differentiated from uptake by using trypsin treatment to degrade non-internalized HSP70. Specificity of HSP70 binding was tested by inhibiting uptake of¹²⁵I-HSP70 with excess unlabeled HSP70 or bovine serum albumin (BSA). The effect of unlabeled exogenous HSP70 on endogenous HSP synthesis was also tested. Exogenous HSP70 increased total cell-associated HSP70 2.9- to 3.6-fold over levels present in unstressed aSMCs. However, <5% of the exogenous HSP70 was trypsin-insensitive, indicating that bound HSP70 was not internalized. Binding of¹²⁵I-HSP70 was inhibited by both unlabeled HSP70 and BSA, implying a non-specific interaction with the plasmalemma. Exogenous HSP70 significantly lowered overall protein synthesis by serum-deprived aSMCs, but it did not specifically inhibit synthesis of endogenous HSPs after heat shock. The results indicate that exogenous HSP70 protects viability of stressed aSMCs through interactions with the cell surface rather than via internalization.     

Association analysis of HSP70A1A haplotypes with heat tolerance in Chinese Holstein cattle

Single-nucleotide polymorphisms (SNPs) in the coding and untranslated regions of heat shock 70 kDa protein 1A (HSP70A1A), an inducible molecular chaperone that is responsible for cellular protection against heat stress, have been reported as being associated with heat tolerance. A fragment of the HSP70A1A gene was amplified in Chinese Holstein cattle and eight novel mutations were found. We performed comprehensive linkage disequilibrium (LD) and haplotype analyses of the eight SNPs of the HSP70A1A gene and examined their involvement in heat resistance in 600 Chinese Holstein cattle. Our results revealed the presence of significant differences between individuals carrying haplotype 1 and those without haplotype 1 for most of the heat-tolerance traits. Haplotype 1 increased the risk of heat stress; however, association analysis of its combination with haplotype 2 showed the lowest rectal temperature and red blood cell K⁺ level, moderate respiratory rate, and the highest red blood cell NKA level, suggesting a heterozygote advantage in the penetration of the phenotype. Protein expression levels in white blood cells among haplotype combinations further confirmed the hypothesis that heterozygotes for haplotypes 1 and 2 are more sensitive to heat stress. We presume that these mutations may be useful in the future as molecular genetic markers to assist selection for heat tolerance in cattle.     

Summer season related heat and nutritional stresses on the adaptive capability of goats based on blood biochemical response and hepatic HSP70 gene expression

A study was conducted to assess the impact of summer season-related heat stress and nutritional stress individually as well as simultaneously on the adaptive capability of goats. Twenty four adult Osmanabadi bucks (average body weight (BW) 16.0 kg) were divided into four groups, C (n = 6; control), HS (n = 6; heat stress), NS (n = 6; nutritional stress) and CS (n = 6; combined stress). The study was conducted for a period of 45 days. The NS and CS bucks were under restricted feed to induce nutritional stress. The HS and CS bucks were exposed to summer heat stress. The animals exhibited different physiological adaptive behavior in the morning and afternoon. Further, the higher plasma cortisol (p < 0.01) and aldosterone (p < 0.05) was recorded in CS group as compared to other groups. The highest degree of degenerative changes and hyperactivity of endocrine cells was recorded in CS group liver and adrenal gland respectively. The higher expression of adrenal and hepatic Heat Shock Protein 70 (HSP70)messenger ribonucleic acid (mRNA) was reported in CS and HS goats respectively. It can be concluded from this study that plasma cortisol and adrenal HSP70 gene expression may be considered as ideal biological markers for combined stresses in Osmanabadi bucks.     

Diminished reproductive potential of male mice in response to selenium‐induced oxidative stress: Involvement of HSP70, HSP70‐2, and MSJ‐1

The oxidative stress imposed by nutritional variations in selenium (Se) has plausible role in reproductive toxicology and affects the reproductive potential. Also, the expression of heat shock proteins (HSPs) is a highly regulated event throughout the process of spermatogenesis and is modulated by stressful stimuli. This prompted us to investigate the possibility that Se‐induced oxidative stress may affect the fertility status by altering the expressions of the constitutive and inducible HSP70 proteins, having crucial role in spermatogenesis. Different Se status‐deficient, adequate, and excess, male Balb/c mice were created by feeding yeast‐based Se‐deficient diet (group I) and deficient diet supplemented with Se as sodium selenite at 0.2 and 1 ppm Se (group II and III) for a period of 8 weeks. After completion of the diet‐feeding schedule, a significant decrease in the Se and glutathione peroxidase (GSH‐Px) levels was observed in the Se‐deficient group (I), whereas Se‐excess group (III) demonstrated an increase. Increased levels of reactive oxygen species, malondialdehyde, and alterations in the redox status in both groups I and III indicated oxidative‐stressed conditions. There was an overall reduced fertility status in mice supplemented with Se‐deficient and Se‐excess diet. The mRNA and protein expression of HSP70 was found to be elevated in these two groups, whereas the expression patterns of HSP70‐2 and MSJ‐1 demonstrated a reverse trend. In vitro CDC2 kinase assay showed reduced kinase activity in group I and group III. These findings suggest that Se‐induced oxidative stress by differentially regulating various HSP70s can affect its downstream factors having crucially important role in differentiation of germ cells and completion of spermatogenesis. Therefore, it can provide an insight into the mechanism(s) by which the oxidative stress–induced reproductive toxicity can lead to increased apoptosis/growth arrest and infertility. This will thus add new dimensions to the molecular mechanism underlying the human male infertility and open new vistas in the development of various chemo‐preventive methods. © 2009 Wiley Periodicals, Inc. J Biochem Mol Toxicol 23:125–136, 2009; Published online in Wiley InterScience ( www.interscience.wiley.com ). DOI 10.1002/jbt.20276     

Secretion of the heat shock proteins HSP70 and HSC70 by baby hamster kidney (BHK‐21) cells

The HSPs (heat‐shock proteins) of the 70‐kDa family, the constitutively expressed HSC70 (cognate 70‐kDa heat‐shock protein) and the stress‐inducible HSP70 (stress‐inducible 70‐kDa heat‐shock protein), have been reported to be actively secreted by various cell types. The mechanisms of the release of these HSPs are obscure, since they possess no consensus secretory signal sequence. We showed that baby hamster kidney (BHK‐21) cells released HSP70 and HSC70 in a serum‐free medium and that this process was the result of an active secretion of HSPs rather than the non‐specific release of the proteins due to cell death. It was found that the secretion of HSP70 and HSC70 is independent of de novo protein synthesis. BFA (Brefeldin A) did not inhibit the basal secretion of HSPs, indicating that the secretion of HSP70 and HSC70 from cells occurs by a non‐classical pathway. Exosomes did not contribute to the secretion of HSP70 and HSC70 by cells. MBC (methyl‐β‐cyclodextrin), a substance that disrupts the lipid raft organization, considerably reduced the secretion of both HSPs, indicating that lipid rafts are involved in the secretion of HSP70 and HSC70 by BHK‐21 cells. The results suggest that HSP70 and HSC70 are actively secreted by BHK‐21 cells in a serum‐free medium through a non‐classical pathway in which lipid rafts play an important role.     

Small heat shock protein p26 associates with nuclear lamins and HSP70 in nuclei and nuclear matrix fractions from stressed cells

The small heat shock/alpha-crystallin protein p26 undergoes nuclear translocation in response to stress in encysted embryos of the brine shrimp Artemia franciscana. About 50% of total p26 translocates to nuclei in embryos treated with heat shock or anoxia, and in embryo homogenates incubated at low pH. Nuclear fractionation shows that the majority of nuclear p26 and a nuclear lamin are associated with the nuclear matrix fraction. To further explore the roles of p26 and other HSPs in stabilizing nuclear matrix proteins (NMPs), nuclear matrices from control, and heat-shocked embryos were disassembled in urea and evaluated by one and two-dimensional (2-D) gel electrophoresis and Western immunoblotting after reassembling. Nuclear lamins were present only in reassembled fractions and, in the case of heat shock, p26 and HSP70 were also present. HSP90 was not detected in any nuclear fraction. Confocal microscopy on isolated nuclei and nuclear matrix preparations from control and heat-shocked embryos showed that the majority of p26 and a nuclear lamin share similar nuclear distributions. The combination of microscopy and fractionation results suggests that p26 and HSP70 play a role in the protection of nuclear lamins within the nuclear matrix.     

The role of the heat shock proteins (HSP70 and sHSP) in the thermotolerance of freshwater amphipods from contrasting habitats

The aim of this study was to clarify the significance of HSP70 and sHSP for thermotolerance in freshwater amphipods. We compared four amphipod species from different freshwater habitats and biogeographical regions (Central Europe vs. Lake Baikal). Test individuals were exposed to thermal stress generated by a water temperature of 25 °C. The thermotolerance of the species, determined by median lethal time (LT50), followed in decreasing order by Gmelinoides fasciatus, Echinogammarus berilloni, Gammarus pulex, Eulimnogammarus verrucosus. HSP70 and sHSP base level concentrations for the species were determined at control (i.e. non-stress) conditions. For HSP70, the base levels were positively correlated to the species' thermotolerances. For sHSP, however, only thermotolerant G. fasciatus showed a high level. Thermal stress at 25 °C water temperature caused a deferred onset of HSP70 and sHSP expression followed by a subsequent offset, delineating a unimodal response curve. The time lag to the expression onset of HSP70 was shorter in the thermosensitive species, compared to thermotolerant ones. Conversely, the time span until the maximum level of HSP70 was variable, not showing a dependence on the thermotolerance properties of the species. The peak concentration in G. pulex was distinctly higher than in the other species, whereas E. verrucosus did not develop a well-defined response maximum at all. In sHSP, the temporal pattern of expression was even more variable than in HSP70. However, the thermosensitive species E. verrucosus showed a time lag of expression onset significantly shorter than the other species and thermotolerant G. fasciatus developed the most pronounced response maximum. Basing on these results, the cellular response to thermal stress in amphipods is more consistently reflected by HSP70, compared to sHSP.     

Modulation of ASK1 expression during overexpression of Trx and HSP70 in stressed fish liver mitochondria

Mitochondrial heat shock protein 70 (mtHSP70) is found to play a primary role in cellular defense against physiological stress like exposure to environmental contaminants and helpful in the maintenance of cellular homeostasis by promoting the cell survival. In the present investigation, the environmental-stress-induced increase in mtHSP70 levels along with the quantification of apoptosis signal regulating kinase 1 (ASK1) and thioredoxin (Trx) were measured in the liver mitochondria of grey mullets (Mugil cephalus) collected from the polluted Ennore estuary and the unpolluted Kovalam estuary for a period of 2 years. The results showed elevated lipid peroxide (LPO) and decreased total antioxidant capacity along with the decrease in mitochondrial viability percentage. Mitochondrial HSP70, ASK1, and Trx levels were increased under this stress condition. A 42% increase in LPO levels and 18% decrease in mitochondrial survivality were observed in the polluted-site fish liver mitochondria when compared to the results of unpolluted estuary. We also report that, under observed oxidative stress condition in Ennore fish samples, the ASK1 levels are only moderately elevated (13% increase). This may be due to mitochondrial-HSP70-induced adaptive tolerance signaling for the activation of Trx (22% increase) which suppresses the ASK1 expression thereby promoting the cell survival that leads to the maintenance of the cellular homeostasis.     

Comparative analysis of differential gene expression of HSP40 and HSP70 family isoforms during heat stress and HIV-1 infection in T-cells

Heat shock proteins (HSPs) are a family of cellular proteins involved in a variety of biological functions including chaperone activity. HSPs are classified based on their molecular weight and each family has several isoforms in eukaryotes. HSP40 is the most diverse family acting as a co-chaperone for the highly conserved HSP70 family. Some of the isoforms are reported to be induced during heat stress. Few studies have also highlighted the diverse role of some isoforms in different stress conditions including viral infections. But till date, no study has comprehensively examined the expression profile of different HSP40 and 70 isoforms in either heat stress or HIV-1 infection, a virus that is responsible for the pandemic of AIDS. In the present study, we have compared the mRNA expression profile of HSP40 and HSP70 isoforms during heat stress and HIV-1 infection in a T-cell line and also validated the HIV-1 stress results in peripheral blood mononuclear cells. In case of HSP70, we observed that three isoforms (HSPA1A, HSPA1B, and HSPA6) are highly upregulated during heat stress, but these isoforms were found to be downregulated during the peak of HIV-1 infection. While in case of HSP40, we found that only DNAJA4, DNAJB1, and DNAJB4 showed significant upregulation during heat stress, whereas in HIV-1 infection, majority of the isoforms were induced significantly. Stress-dependent differential expression observed here indicates that different HSP40 and HSP70 isoforms may have specific roles during HIV-1 infection and thus could be important for future studies.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12192-020-01185-y.     

植物热激蛋白70的分子作用机理及其利用研究进展

热激蛋白70（heat shock protein 70,HSP70）广泛参与胁迫环境的响应,在诱发人体肿瘤细胞凋亡,增强肿瘤的免疫原性中起着重要作用。然而,植物HSP70的生理功能研究起步较晚,最近的研究发现植物HSP70在细胞内主要参与新生肽的折叠与成熟、损伤蛋白的降解和蛋白运输;植物HSP70在非生物胁迫环境的应答、抗病性及植物发育中起着重要作用。本文从分子生物学角度,系统综述了植物HSP70分子作用机理研究的进展,以及在提高植物抗逆性方面的作用,以期为基因工程方法改良作物抗性提供参考。     

朱砂叶螨热激蛋白HSP70基因TCHSP70-4的克隆与表达

为了研究朱砂叶螨Tetranychus cinnabarinus(Boisduval)热激蛋白HSP70的表达与其适应高温和低温胁迫的关系,我们利用物种的同源性及RACE 技术,获得朱砂叶螨热激蛋白HSP70基因1个,命名为TCHSP70-4（GenBank 登录号为EU977182）。该基因全长2 182 bp,包含1 959 bp的开放阅读框,编码653个氨基酸,理论分子量为70.9 kDa,等电点为5.4,含有HSP70家族高度保守的基序。运用real-time PCR分析冷激（4℃）和热激（40℃）1 h后TCHSP70-4在朱砂叶螨体内的表达量。结果显示冷激后TCHSP70-4表达量明显下降,而热激后TCHSP70-4表达量却明显上升。这些结果一方面表明该基因属于诱导型HSP70基因,另一方面揭示了朱砂叶螨分别受到冷和热胁迫后体内TCHSP70-4的表达及所起的保护作用是不同的。     

谢氏宽漠王HSP70基因cDNA片段的克隆及热激条件下的表达

谢氏宽漠王Mantichorula semenowi Reitter是一种沙漠指示性甲虫。本研究由该种甲虫体内克隆到两种不同的HSP70基因片段,分别为MsHSP70和MsHSC70。同源性发现表明这两个基因片段与已报道的其他昆虫的热休克蛋白核苷酸序列高度同源。半定量RT-PCR分析显示：经42℃热激1 h 后立即诱导MsHSP70表达至最高峰;在恢复到室温的1～4 h 内MsHSP70表达量逐渐降低,但仍然高于未热激对照组。而MsHSC70在42℃热激1 h后表达受到抑制,但在恢复2 h和4 h时有少量的表达,分别仅为未热激对照组的0.25和0.28倍。结果提示MsHSP70和MsHSC70在保护细胞方面具有不同的作用。本实验结果为谢氏宽漠王在极端的沙漠环境胁迫下的抗逆适应性研究提供了理论基础。     

Coordinate and non-coordinate expression of the stress 70 family and other molecular chaperones at high and low temperature in spinach and tomato

Stress 70 molecular chaperones are found in all the major subcellular compartments of plant cells, and they are encoded by a multigene family. Twelve members of this family have been identified in spinach. The expression of the stress 70 molecular chaperones in response to heat shock is well-known and it appears that low temperature exposure can also stimulate their expression. However, it has been difficult to determine which member(s) of the family are specifically responsive to low temperature. This study was initiated to determine the levels of expression of the stress 70 family members and other selected chaperones in response to high and low temperature exposure. During heat shock of spinach, of the 10 stress 70 family members that were examined, all 10 showed increased RNA levels after one hour, and all showed down-regulation at longer durations of high temperature exposure. However, the response to low temperature was quite variable and complex. Some members were induced, some were transiently up-regulated, while others showed sustained up-regulation at a low non-freezing temperature. In comparison, the entirety of the molecular chaperone expression response of cold-sensitive tomato at the same low non-freezing temperature was even more dramatic with 11 of 15 molecular chaperones tested exhibiting elevated expression. The increased chaperone expression is consistent with the hypothesis that the biogenesis or stability of some proteins is compromised at low non-freezing temperatures. In contrast, mild freezing sufficient to cause injury of spinach did not materially activate chaperone expression.     

Effect of sericin supplementation on heat shock protein 70 (HSP70) expression,redox status and post thaw semen quality in goat

Cryopreservation results in substantial deterioration of heat shock protein 70 (HSP70) and ultra-structural changes in sperm organelles, resulting in a marked reduction in post-thaw semen quality. The present study was aimed to explicate the effect of sericin supplementation on expression profile of HSP70, redox status and post-thaw semen quality in Barbari goat. Five Barbari bucks were used to collect thirty semen ejaculates by using artificial vagina and each ejaculate was divided into three aliquots to which sericin was supplemented at 0% (Control), 0.25% (T1) and 0.50% (T2). Further, extended semen samples were equilibrated followed by their cryopreservation. Post-thaw semen characteristics, redox status of seminal plasma, enzyme leakage and HSP70 gene/protein expression in spermatozoa were assessed in all the groups. Per cent progressive motile spermatozoa, spermatozoa having intact plasma membrane (HOST + ve) and intact acrosomes in post-thaw spermatozoa were significantly (p < 0.01) higher in T1 and T2 as compared to control. A significant (p < 0.01) reduction in abnormal spermatozoa was found in T1 as compared to T2. Sericin supplementation significantly (p < 0.05) improved the antioxidative status (SOD, GST, CAT), reduced lipid peroxidation (MDA) and also prevented enzyme (ALT, LDH) leakage as compared to control samples. qRT-PCR results revealed that HSP70 mRNA expression was significantly (p < 0.01) upregulated in T1 and T2 group as compared to control. The positive effect of sericin on expression of HSP70 was further confirmed by immunoblotting followed by densitometry revealing higher expression in T1 and T2 compared to control. Inclusion of 0.25% w/v sericin in semen extender ameliorated the post-thaw semen quality by improving antioxidative status and minimizing the leakage of intracellular enzymes. Sericin supplementation had a beneficial effect on HSP70/HSP70 mRNA expression either by induction or by protection of HSP70/HSP70 mRNA as evident from the gene expression and immunoblotting studies.     

Effects of high ambient temperature on respiration rate, rectal temperature, fetal development and thyrold gland activity in tropical and temperate breeds of sheep

The effects of high ambient temperatures on rectal temperature (RT), respiration rate (RR), fetal development and serum thyroxine (T(4)) concentrations were stuaied in two experiments involving 35 ewes and 26 lambs from the following ewe groups: 1) Barbados Blackbelly (B), a tropical breed; 2) Dorset (D), a temperate breed; and 3) Blackbelly x Dorset crosses (BxD). Data were obtained on four B, five D and five BxD ewes exhibiting estrus during the summer (Exp. 1). In Exp. 2, eight B, seven D and six BxD ewes were maintained in two environmental chambers (cool, 22.2C; hot, 33.8C) from day 125 of gestation to seven days before the expected lambing date for each breed group (D and BxD, 140+/-1; B, 144+/-1 day of gestation). The B and BxD ewes were more heat-tolerant than D ewes as measured by significantly lower RT and RR in each experiment. Mean lamb birth weight, crown-rump length, number of functional uterine caruncles and caruncle weight and size did not vary significantly among breed groups or temperature chamber (Exp. 2), and there was no indication that the high temperature imposed caused fetal dwarfing in lambs removed from the uterus at a standard age of seven days before expected parturition. Serum T(4) varied markedly among breed groups (P<0.05) in each experiment with B ewes having the lowest and BxD ewes the highest concentration. In Exp. 1, follicular stage T(4) concentrations in B and BxD ewes were lower (P<0.02) than those during the luteal stage of the estrous cycle. The decrease in D ewes was not significant. High ambient temperature (Exp. 2) depressed T(4) levels in D ewes (P<0.05) and also depressed the pituitary-thyroid response to thyrotropin releasing hormone in D lambs. Such was not the case in B and BxD ewes and their lambs.