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Biomedical subjects

S M Rhind

Publications and source records attributed to S M Rhind.

At least 37 records · Page 2Linked to original sources

Endocrine disrupting compounds and farm animals: their properties, actions and routes of exposure.

Endocrine disrupting compounds (EDC) comprise a diverse group of compounds of anthropogenic origin, including organochlorine pesticides, alkyl phenols, phthalates, polychlorinated biphenyls (PCB), dioxins and polybrominated diphenyl ethers. EDC are generally present in the environment at low concentrations but they are ubiquitous and persistent and, although environmental concentrations are low, they appear to exert a range of adverse effects on animals of many species, including humans. Their effects include disruption of reproductive function and of the immune system and they can be carcinogenic. Animals may be exposed to relatively high concentrations of EDC because they persist in the environment and when ingested, they may be concentrated in fat tissue and released when the fat is mobilised during pregnancy or lactation, thus exposing, to relatively high concentrations, embryos and neonates. These stages of development are particularly susceptible to EDC effects. Very little is known of EDC body burdens in domestic animal species and particularly in those exposed to unpolluted environments. EDC concentrations in soils and plant material and their rates of ingestion and metabolism have been little studied but it is concluded that there is a potential risk of significant bioaccumulation and of associated effects on the health and reproductive capacity of domestic animals and of humans consuming animal products.

Animal Feed↗

Maternal undernutrition alters triiodothyronine concentrations and pituitary response to GnRH in fetal sheep.

The aims of this study were to determine which hormones may have a role in the expression of maternal undernutrition effects on reproductive function, in both the developing fetus and the adult offspring. This was undertaken by measuring the effects of long-term maternal undernutrition on metabolic hormone profiles and pituitary responses to single doses of GnRH and GH-releasing factor (GRF) in fetal sheep. From mating, groups of ewes were fed rations providing either 100% (HIGH) or 50% (LOW) of estimated metabolisable energy requirements for pregnancy throughout the experiment until slaughter at approximately 119 days of gestation. Fetal and maternal blood samples were collected from 113 until 119 days of gestation, via carotid and jugular catheters respectively, and assayed for insulin, IGF-I, GH, thyroxine and triiodothyronine (T(3)). Undernutrition had no effects on fetal weight, fetal gonad weight of either sex, fetal insulin or IGF-I concentrations. Male LOW fetuses exhibited a significantly attenuated response (P<0.05) to a bolus challenge of GnRH compared with HIGH fetuses. Basal fetal GH concentrations and the response to exogenous GRF were similar in both treatment groups, although LOW fetuses exhibited more secretory episodes (P<0.01). Mean T(3) concentrations were significantly lower in both the maternal (P<0.01) and fetal (P<0.05) plasma of LOW animals compared with HIGH animals. It is concluded that pituitary function was altered in fetal males and could influence male reproductive development. On the other hand, in female sheep, fetal gonadal abnormalities and reductions in reproductive capacity in adult life which are associated with fetal undernutrition are unlikely to be attributable to altered pituitary function. Additionally, these studies raise the possibility that thyroid hormones may have a role in the expression of maternal undernutrition effects on fetal development.

Analysis of Variance↗

Contrasting effects of different levels of food intake and adiposity on LH secretion and hypothalamic gene expression in sheep.

Body reserves (long-term) and food intake (short-term) both contribute nutritional feedback to the hypothalamus. Reproductive neuroendocrine output (GnRH/LH) is stimulated by increased food intake and not by high adiposity in sheep, but it is unknown whether appetite-regulating hypothalamic neurons show this differential response. Castrated male sheep (Scottish Blackface) with oestradiol implants were studied in two 4 week experiments. In Experiment 1, sheep were fed to maintain the initial body condition (BC) score of 2.0+/-0.00 (lower BC (LBC), n=7) or 2.9+/-0.09 (higher BC (HBC), n=9), and liveweight of 43+/-1.1 and 59+/-1.6 kg respectively. LBC and HBC sheep had similar mean plasma LH concentration, pulse frequency and amplitude, but HBC animals had higher mean plasma concentrations of insulin (P<0.01), leptin (P<0.01) and glucose (P<0.01). Gene expression (measured by in situ hybridisation) in the hypothalamic arcuate nucleus (ARC) was higher in LBC than HBC sheep for neuropeptide Y (NPY; 486% of HBC, P<0.01), agouti-related peptide (AGRP; 467%, P<0.05) and leptin receptor (OB-Rb; 141%, P<0.05), but lower for cocaine- and amphetamine-regulated transcript (CART; 92%, P<0.05) and similar between groups for pro-opiomelanocortin (POMC). In Experiment 2, sheep with initial mean BC score 2.4+/-0.03 and liveweight 55+/-0.8 kg were fed a liveweight-maintenance ration (low intake, LI, n=7) while sheep with initial mean BC score 2.0+/-0.03 and liveweight 43+/-1.4 kg were fed freely so that BC score increased to 2.5+/-0.00 and liveweight increased to 54+/-1.4 kg (high intake, HI, n=9). Compared with LI, HI sheep had higher mean plasma LH (P<0.05), baseline LH (P<0.01) and pulse amplitude (P<0.01) and showed a trend towards higher pulse frequency. Although there were no differences in final mean plasma concentrations, there were significant increases over time in mean concentrations of insulin (P<0.001), leptin (P<0.05) and glucose (P<0.001) in HI sheep. Gene expression for AGRP in the ARC was higher in HI than LI animals (453% of LI; P<0.05), but expression levels were similar for NPY, OB-Rb, CART and POMC. Thus, the hypothalamus shows differential responses to steady-state adiposity as opposed to an increase in food intake, in terms of both reproductive neuroendocrine activity and hypothalamic appetite-regulating pathways. Differences in hypothalamic gene expression were largely consistent with contemporary levels of systemic leptin and insulin feedback; however, increased nutritional feedback was stimulatory to GnRH/LH whereas constant high feedback was not. The hypothalamus therefore has the ability to retain a nutritional memory that can influence subsequent responses.

Adipose Tissue↗

Impact of maternal nutrition during pregnancy on pituitary gonadotrophin gene expression and ovarian development in growth-restricted and normally grown late gestation sheep fetuses.

The influence of maternal nutrition during pregnancy on anterior pituitary gonadotrophin gene expression and ovarian development in sheep fetuses during late gestation was investigated. Embryos recovered from superovulated adult ewes that had been inseminated by a single sire were transferred, singly, into the uteri of adolescent recipients. After embryo transfer, adolescent ewes were offered a high or moderate amount of a complete diet. Pregnancies were terminated at day 131 +/- 0.6 of gestation and the fetal brain, anterior pituitary gland and gonads were collected. Gonadotrophin gene expression (LHbeta and FSHbeta subunits) in the fetal pituitary gland was examined using in situ hybridization. Ovarian follicular development was quantified in haematoxylin- and eosin-stained ovarian sections embedded in paraffin wax. Six dams that were offered a high nutrient intake carried normal-sized fetuses (weight within +/- 2 SD of mean weight for control fetuses from dams fed a moderate level of complete diet) and 13 dams carried growth-restricted fetuses (weight <or= +/- 2 SD of mean weight for control fetuses from dams fed a moderate level of complete diet). Mean placental masses in these groups were 354 +/- 24.5 and 230 +/- 21.1 g, respectively, compared with 442 +/- 54.3 g in the dams that were offered a moderate nutrient intake (n = 6). Growth-restricted fetuses from dams offered a high nutrient intake showed higher pituitary LHbeta mRNA expression (P < 0.05) than normal-sized fetuses from dams offered a moderate nutrient intake (252 +/- 21.6 and 172 +/- 23.6 nCi g(-1), respectively). FSHbeta mRNA expression was not influenced by growth status. Fewer follicles (primarily in the resting pool) were observed in the ovaries of both growth-restricted (P < 0.002) and normal-sized fetuses from dams offered a high nutrient intake (P < 0.01) compared with normal-sized fetuses from dams offered a moderate nutrient intake. Irrespective of nutritional treatment, the total number of follicles was positively associated with placental mass (P < 0.01). Thus, a high maternal nutrient intake during adolescent pregnancy had a negative influence on ovarian follicular development in fetuses as determined during late gestation.

Animal Nutritional Physiological Phenomena↗

Effect of maternal undernutrition on fetal testicular steroidogenesis during the CNS androgen-responsive period in male sheep fetuses.

The aim of this study was to determine the effects of maternal undernutrition, applied during physiologically relevant stages of development of the reproductive system, on reproductive development in male sheep fetuses. Groups of ewes (n = 11-19) were fed rations providing either 100% (high; H) or 50% (low; L) of metabolizable energy requirements for live weight maintenance during selected 'windows', bounded by days 0, 30, 50, 65 and 110 after mating. Ewes of control groups (HH (Expts 1 and 2) and HHH (Expt 3)) were fed the H ration from mating until they were killed at day 50 (Expt 1), day 65 (Expt 2) or day 110 (Expt 3) of gestation, whereas ewes of other groups were fed the L ration for the periods days 0-30 of gestation (LH and LHH), days 31-50 or days 31-65 of gestation (HL and HLH), days 65-110 of gestation (HHL), or day 0 to day 50, day 65 or day 110 of gestation (LL and LLL) when the animals were killed. At day 50 of gestation, there was no effect of nutritional treatment on mean fetal mass or fetal testicular mass, but there was increased expression of mRNA for steroidogenic acute regulatory protein (StAR) in the testes of LL animals (P < 0.05) compared with HH controls. Compared with HH animals, the mean plasma testosterone concentrations of LL fetuses tended to be higher, but this result did not reach significance. At day 65 of gestation there were no significant differences between treatments in mean fetal masses, testicular masses, mean plasma testosterone concentrations or StAR mRNA content. At day 110 of gestation, fetal masses in the LLL group were lower (P < 0.01) than those of control fetuses, although no differences in testicular size or fetal plasma testosterone concentrations were recorded. It is concluded that the effects of undernutrition on reproductive development of male sheep fetuses are dependent on the timing of the period of undernutrition.

Analysis of Variance↗

The role of cholecystokinin in the expression of seasonal variation in the feed intake and eating pattern of red deer (Cervus elaphus).

The role of cholecystokinin (CCK) in the expression of seasonal variation in feed intake and feeding activity in ruminants was investigated by active immunisation of castrated male red deer (Cervus elaphus). In April, animals of two groups (five animals per group) were immunised against either CCK or vehicle solution only and booster injections were administered at 2-month intervals for the following year. Measurements were conducted for a period of more than a year from July. There were no significant effects of immunisation on mean daily food intake. However, there was a significant interaction (P<.01) between immunisation and month (season), with respect to rate of feed ingestion during meals, with animals immunised against CCK exhibiting higher mean rates of ingestion during October to May but lower mean rates during June to September. It is concluded that systemic CCK has a role in the expression of seasonal variation in the rate of feed ingestion during meals in ruminants and that this, in turn, may affect the pattern of seasonal change in daily feed intake. Since there was no evidence of differences with treatment in profiles of insulin, growth hormone (GH), thyroxine (T(4)), triiodothyronine (T(3)), insulin-like growth factor-1 (IGF-1) or prolactin, it is unlikely that this effect is expressed through changes in the profiles of these hormones.

Animals↗

Natural paratuberculosis infection in rabbits in Scotland.

Natural paratuberculosis infection of rabbits (Oryctolagus cuniculus) was recently diagnosed in Scotland, and an investigation into the pathology of the disease in wild rabbits is reported in this paper. Evidence of Mycobacterium avium subsp. paratuberculosis (M.a. paratuberculosis) infection was detected in 22% of 110 rabbits; the organism was cultured from 17 of 110 rabbits, and histopathological lesions consistent with M.a. paratuberculosis infection were noted in 18 of 98 rabbits examined. No macroscopical lesions suggestive of M.a. paratuberculosis infection were observed. The histopathological lesions were either severe or mild. Severe lesions consisted of extensive macrophage granulomata and numerous giant cells, with many intracellular acid-fast bacteria in the small intestine. For the examination of formalin-fixed, paraffin wax-embedded tissues, neither immunohistochemistry nor the polymerase chain reaction was as sensitive a method of diagnosis as histopathology.

Animals↗

Experimental paratuberculosis in calves following inoculation with a rabbit isolate of Mycobacterium avium subsp. paratuberculosis.

The role of wildlife species in the epidemiology of paratuberculosis has been the subject of increased research efforts following the discovery of natural paratuberculosis in free-living rabbits from farms in east Scotland. This paper describes the experimental inoculation of young calves with an isolate of Mycobacterium avium subsp. paratuberculosis recovered from a free-living rabbit. After a 6-month incubation period, all eight calves inoculated with the rabbit isolate had developed histopathological and/or microbiological evidence of M. avium subsp. paratuberculosis infection. Similar results were obtained from a group of calves infected with a bovine isolate of M. avium subsp. paratuberculosis. The virulence of the rabbit isolate for calves demonstrated in this study suggests that rabbits are capable of passing paratuberculosis to domestic ruminants and that wildlife reservoirs of M. avium subsp. paratuberculosis should therefore be considered when formulating control plans for the disease.

Animals↗

Hypothalamic control of photoperiod-induced cycles in food intake, body weight, and metabolic hormones in rams.

This study used a hypothalamo-pituitary disconnected (HPD) sheep model to investigate the central regulation of long-term cycles in voluntary food intake (VFI) and body weight (BW). VFI, BW, and circulating concentrations of metabolic hormones [alpha-melanocyte-stimulating hormone (alpha-MSH), insulin-like growth factor-1 (IGF-1), insulin, and leptin] were measured in HPD and control Soay rams exposed to alternating 16 weekly periods of long and short days for 80 wk. In the controls, the physiology was cyclical with a 32-wk periodicity corresponding to the lighting regimen. VFI and BW increased under long days to a maximum early into short days, and there were associated increases in blood concentrations of alpha-MSH, insulin, and leptin. In the HPD rams, there were no significant photoperiod-induced changes in any of the parameters. VFI increased after surgery for 8 wk and then gradually declined, although BW increased progressively and the HPD rams became obese. Concentrations of alpha-MSH, insulin, and leptin in peripheral blood were permanently increased (>200%), and levels of IGF-1 decreased (<55%). The HPD lesion effectively destroyed the entire median eminence [no nerve terminals immunostained for tyrosine hydroxylase (TH) and gonadotropin-releasing hormone] and the adjacent arcuate nucleus (no perikarya immunostained for proopiomelanocortin or TH, and no cells expressed neuropeptide Y mRNA). The results support the conclusion that arcuate hypothalamic systems generate long-term rhythms in VFI, BW, and energy balance.

Adaptation, Physiological↗

CD1--the pathology perspective.

CD1 molecules are a family of cell surface-associated glycoproteins now recognized as having a role in antigen presentation. These glycoproteins are distinct from yet have some similarities to classical major histocompatibility complex class I and class II molecules. The role of these molecules has been studied in detail over recent years, with an explosion of interest following the demonstration that they can present nonprotein antigens to certain subpopulations of T cells. The purpose of this paper is to provide an overview of current knowledge of the function of the CD1 family with specific emphasis on the potential role in the pathogenesis of certain diseases. Although much of the current research in this field has inevitably concentrated on mice and humans, this work also has potential significance for veterinary species.

Animals↗

Effects of nutrition and environmental factors on the fetal programming of the reproductive axis.

Research from a wide range of scientific disciplines has shown that the reproductive performance of animals in adult life is determined, in part, by a variety of extraneous influences acting at different stages of development from before conception until after birth. These effects are probably mediated through changes in the hypothalamic-pituitary and gonadal axes but the physiological system that is affected depends on the stage of development at which the influence is applied. The physiological mechanisms through which environmental influences are transmitted to the target organs are, in many cases, complex and poorly understood. Gonadotrophins seem to play a pivotal role in the development of the fetal testis, although effects of environmental influences on GnRH secretion have yet to be demonstrated. Other studies have shown that, at earlier stages of fetal development, the normal ontogeny of gonadal development and function can be disrupted by undernutrition or the influence of endocrine-disrupting compounds. Specifically, in female fetuses, the onset of meiosis is delayed, whereas, in male fetuses, testosterone synthesis is increased as a result of enhanced testicular steroidogenic enzyme activity. Although reproductive performance is clearly influenced by prenatal factors, much further work is required to identify the relationships between developmental abnormalities and adult reproductive function. Work is also required to elucidate further the critical windows in development and the mechanisms by which environmental factors affect the reproductive organs of developing offspring.

Animals↗

Influence of placentally mediated fetal growth restriction on the onset of puberty in male and female lambs.

The onset of puberty in prenatally growth-restricted versus normally grown lambs of both sexes, born in April and housed under natural photoperiod, was examined. Singleton pregnancies were established and adolescent ewes were offered a high or moderate nutrient intake throughout gestation. Placental mass was reduced (P < 0.001) in high compared with moderate intake dams and resulted in the birth of growth-restricted and normal birth weight offspring, respectively. At birth, female lambs weighed 3.43 kg versus 5.03 kg (P < 0.001; n = 14 per group) and male lambs weighed 2.75 kg versus 5.18 kg (P < 0.001; n = 7 per group) in growth-restricted and normal birth weight groups, respectively. Lambs suckled for 12 weeks and thereafter were fed ad libitum until week 43 of age. Growth-restricted lambs had lower preweaning live weight gains and this difference was more pronounced in male (P < 0.05) than in female lambs (P = 0.07). Thereafter, live weight remained lower (P < 0.05) in growth-restricted than in normally grown lambs of both sexes until week 25 of age. In females, the time of onset of puberty was similar in the two groups. All females ovulated and there were no differences in the number of ovarian cycles recorded or in the incidence of aberrant ovarian function. In males, testosterone concentrations and testicular volume were lower in growth-restricted compared with normally developed lambs from birth until weeks 28 and 35 of age, respectively (P < 0.05). The seasonal increase in plasma testosterone concentrations occurred later in growth-restricted than in normally developed lambs (P < 0.01) but the timing of maximum peak concentrations was similar. Peak testosterone concentrations were lower (P < 0.05) in growth-restricted than in normal male lambs.

Aging↗

Effect of maternal undernutrition during pregnancy on early ovarian development and subsequent follicular development in sheep fetuses.

Gonad development in female sheep fetuses is thought to occur in a number of key stages. The aim of this study was to determine the effects of maternal undernutrition, applied at one or more of these critical stages, on fetal ovarian development. Groups of ewes (n = 11-19) were fed rations providing either 100% (high; H) or 50% (low; L) of energy requirements for live weight maintenance during selected 'windows' during gestation. Control ewes (HH and HHH) were fed the H ration from mating until they were killed at days 50, 65 (HH) or 110 (HHH) of gestation, whereas ewes of other groups were fed the L ration for the periods between day 0 and day 30 of gestation (LH and LHH), day 31 and day 50 or 65 of gestation (HL and HLH), day 65 and day 110 of gestation (HHL) or day 0 of gestation until the animals were killed (LL and LLL). At day 50 of gestation, there was no effect of nutritional treatment on mean fetal mass but compared with HH animals, mean fetal ovarian mass was significantly lower in HL (P < 0.05) and LL (P < 0.001) animals. At day 65 of gestation, there were significantly fewer germ cells (P < 0.05) at the resting, diplotene stage of initial meiosis in LL animals than there were in HH animals, indicating delayed germ cell maturation and onset of meiosis. Qualitative assessment of proliferative cell nuclear antigen immunostaining indicated that, at day 50 of gestation, staining was located predominantly in the germ cells, whereas by day 65 of gestation, staining was confined predominantly to somatic cells. Undernutrition in each one of these windows was associated with delayed ovarian follicular development (P < 0.05-0.001) as measured by development of the granulosa cell layer at day 110 of gestation. This study demonstrates that undernutrition before and during folliculogenesis can delay fetal follicular development.

Analysis of Variance↗

Modulation of gammadelta T cells and CD1 in Mycobacterium avium subsp. paratuberculosis infection.

M.a. paratuberculosis is the causal agent of paratuberculosis (Johne's disease). Recent work has suggested that gammadelta T cells may play an important role in the early immunological response to mycobacterial diseases, and that CD1 may act as a non-classical MHC molecule in antigen presentation to these gammadelta T cells. Experimental infection of neonatal lambs with M.a. paratuberculosis was used to investigate the changes in gammadelta T cells and CD1 molecules in the gut associated lymphoid tissue 4 weeks after inoculation. Immunohistochemistry was used to label the gammadelta lymphocytes and CD1 molecules. An increase in the number of gammadelta T cells was noted in both the jejunal and ileal Peyer's patches in the gut of infected lambs, but no statistically significant change was found in the mesenteric lymph nodes. There were no obvious changes in the CD1 molecules in any tissue. This work suggests that gammadelta T cells may play a role in the initial immunological events of paratuberculosis infection.

Animals↗

Sheep CD1 genes and proteins.

Interest in CD1 genes and proteins was initially stimulated by their close evolutionary and structural relationship to MHC class I molecules. The demonstration that CD1b and c molecules present novel non-peptide antigens to T-cells and play a role in protection against mycobacterial infection then focused attention on the functional role of CD1 proteins. Sheep possess at least seven CD1 genes, including CD1B, D and E, which is the most complex genetic arrangement identified so far in any animal. OvCD1B consists of at least three distinct genes, with the probability of limited polymorphism and the existence of splice variants. Most anti-sheep CD1-specific monoclonal antibodies react with OvCDlb and phenotypic and immunochemical data suggests the existence of two variants. CD1D genes have been identified in all species studied, suggesting a conserved role for CDld proteins across mammalian species. Presumptive evidence for the existence of OvCDIE has been obtained by NH2-terminal sequencing of protein precipitated by the mAb 20.27 (SBU-T6). Confirmatory evidence from gene cloning experiments is currently being sought. Collectively, these factors make the sheep CD1 family a highly relevant model for investigating the in vivo role of CD1 molecules. In this survey, the properties of monoclonal antibodies specific for sheep CD1, the cellular distribution and physicochemical characteristics of sheep CD1 molecules and the current state of knowledge on sheep CD1 genetics are reviewed.

Amino Acid Sequence↗

Differential expression of ovine CD1.

was examined using a combination of reverse transcription-polymerase chain reaction (RT-PCR) with sequence-specific primers and Northern and in situ hybridization techniques. The aim of the study was to establish the patterns of CD1 expression at the molecular level and address questions posed by previous studies in other species regarding expression patterns of CD1. A 'pan-CD1' probe based on the exon 4 (alpha 3) region was used in addition to isotype-specific probes for SCD1B (the exon 3 region of clone SCD1B42) and SCD1D (the exon 3 region of clone SCD1D). Widespread expression of CD1 (including thymus, peripheral blood lymphocytes, lung and intestine) was identified using both the exon 4 and SCD1D probe. SCD1B expression was more restricted, being identified in equivalent levels only in the thymus and in scattered populations of dendritic cells. These results highlight the difference in expression patterns between group 1 and group 2 CD1 family members and establish SCD1D as the CD1 family member with the widest pattern of expression, consistent with a differential role for the different CD1 family members.

Animals↗

Type II and type III monodeiodinase activities in the skin of untreated and propylthiouracil-treated cashmere goats.

The presence or absence of types I, II and III iodothyronine monodeiodinase enzymes (MDI, MDII and MDIII) and their levels of activity in the skin of goats, which were orally dosed for 60 days with 0, 1.1, 2.2, 4.4, 8.8, 17.5, or 35 mg(-1)kg liveweight day(-1)of the anti-thyroid, enzyme-inhibiting drug, propylthiouracil (PTU), were determined. Contrary to our earlier report that PTU did not influence skin MDII activity, the currect more thorough investigation (in terms of numbers of observations and the efficiency of the enzyme extraction procedure) indicated that doses of 1.1.to 17.5 mg kg(-1)liveweight induced a 2 to 3 fold increase (P = 0.01) in MDII activity. However, in three of the four goats treated with 35 mg kg(-1)group, activity was similar to that of control animals. There were no significant differences between treatments in MDIII activity but there was a trend towards lower levels of activity in the goats dosed with 17.5 and 35 mg kg(-1). It is concluded that there is significant MDII and MDIII activity in the skin of goats and that although there is none of the PTU -sensitive MDI enzyme, synthesis of T3 within the skin could nevertheless be modified through increases in MDII activity induced by lower T4 concentrations in the circulation caused by PTU. Changes in pattern of fibre moult induced by treatment with low doses of MD-inhibiting drugs may therefore be achieved through this effect. Since MDII and MDIII enzyme activity may be reduced by high doses of PTU, prolonged treatment with high doses of PTU may have adverse effects on skin tissue.

Animals↗

Myasthenia gravis associated with cutaneous lymphoma in a dog.

A middle-aged golden retriever presenting with exercise intolerance and multifocal cutaneous nodules was diagnosed as having non-epitheliotropic cutaneous lymphoma with concurrent myasthenia gravis. Treatment with corticosteroids induced short-term remission of the lymphoma and alleviation of myasthenic signs.

Adrenal Cortex Hormones↗