[Music] [Music] at addiso we cherish the work we carry out with animals and thanks to us chickens and pigs grow with less feed and in a healthier way cows produce milk in greater quantity and quality for a longer period and fish and shrimp now have fewer diseases and grow faster these animals contribute to feed 8 billion people and have enabled us to become a global leader in the production of nutritional solutions for animal feed one of the leaders in methionine an essential amino acid and protected methionine for ruminants one of the key players in the vitamins and specialty products designed to foster animal health and welfare our story began in france with alimote libre in commentary in 1939 then in 1971 the company is acquired by hon pulank becoming its animal feed division in 2002 acquired by an investment fund and renamed adeseo the company regains its independence and in 2006 we joined the blue star group thus asserting opposition in asia and china especially adeseo is a unique company in many aspects our routes are in france but our products come from europe and china and 95 of them are exported to over 110 countries and have more than 3 900 customers worldwide we are a medium-sized company with around 1.5 billion euros turnover and almost 2 200 people working for our deseo our uniqueness also comes from our investments in both industry and research guaranteeing a competitive and innovative product and service offering 1.7 billion euros have been invested since 2007. our success is also due to our dedication to essential values team spirit creativity integrity commitment and results over the years our company has changed developed and grown putting safety and sustainability as top priorities safety of our employees safety of our plants and of our research centers safety of our products we never do any compromise with safety aware of present and future economic environmental and social issues adesayo has developed a growth strategy that is clearly in line with the un's sustainable development goals building on its strengths and successes adesayo strives to be a partner of choice in animal nutrition with women and men who make the difference every day to feed the planet in a high quality affordable safe and sustainable way hello this is dr brian sloan from addiso hello and welcome to today's webinar how heat stress impacts progeny and how nutrients impact heat stress addiso is well known for hosting a wine and cheese reception on the tuesday evening of the american dairy science association's annual meeting this year the adsa meeting is virtual as it was last year so once again our reception is taking the form of a webinar today we continue our legacy of forward-looking and thought-provoking presentations our goal is to prompt discussions and if the past is any predictor of the future i'm sure we will have a lively discussion this evening after the presentations today's topic is how heat stress impacts progeny and how nutrients may be able to mitigate short and long-term effects of heat stress dr jimena laporta an assistant professor in the department of animal and dairy science at the university of wisconsin at madison will join us first originally from uruguay ximena received her bachelor's in biology and masters in animal science in 2014 she earned her phd in dairy science from the university of wisconsin-madison and moved to the university of florida as a faculty member after five years she returned to madison as an assistant professor her program integrates mammary gland physiology with advances in management genetics and nutrition to overcome the challenges daily cattle face across their lifetime including increased susceptibility to the environment metabolic disorders and immune suppression her research incorporates state-of-the-art molecular techniques to investigate how autocrine systemic and environmental factors affect the regulatory of mammary gland development and function as well as no synthesis and composition she also investigates the underlying molecular mechanisms by which prenatal and postnatal stressors contribute to the programming of offspring's future potential she will make significant connections between the cow that is in advance pregnancy during heat stress her offspring and yes even future generations once we understand the situation and the challenge we hear from dr juan lohr who is a professor in the department of animal science at the university of illinois juan's research focus is on nutritional and physiological genomics during lactation and growth in cattle juan earned his bachelor's at the university of california davis for his master's on phd he moved to virginia polytechnic institute and state university in blacksburg he then held a postdoctoral research associate professor position in france before moving to the university of illinois as a postdoctoral research associate at illinois he went on to become a full professor in 2018 his research program focuses on the study of how nutrition physiological state and their interactions affect tissue function and metabolic adaptions through alterations in the transcriptome he heavily relies upon cutting-edge technologies to make this possible you find the integration of resulting data sets at the core of the university's systems biology efforts in nutritional sciences along the way juan has managed to publish numerous book chapters and journal articles so we've asked one to supply his in-depth understanding of nutrition to the challenge jimena outlines this should give you some practical guidance on how heat stress impacts progeny and how nutrients impact heat stress and now without further ado let's welcome dr jimena laporta from the university of wisconsin at madison amena the floor is yours hello everyone i would like to start by thanking alessio for inviting me for giving me this opportunity to share my research today and for you to join in this webinar today i will be focusing on the first part of this webinar series how heat stress impacts the progeny all right so it's no news that our planet is getting warmer in fact according to uh the nasa in 200 years of the surface temperature of the globe increased to fahrenheit and the two warmest years were in 2016 and just last year in 2020 so heat stress is one of the largest challenges affecting dairy cows all across the world even in temperate regions and the us is no exception so his stress is typically associated with southern stern states such as florida georgia texas louisiana which experience more than 150 days of heat stress but if you look at this map here except for california and texas the other five states with the highest population of cows in the us such as pennsylvania wisconsin new york idaho minnesota they are all located in northern regions and those experience an average 50 50 days of heat stress i know that this may not seem like a lot but when you think about the number of cows that are exposed to that short relatively short period of time the economic consequences of heat stress even during this time could be significant so regardless our individual efforts to our collective efforts to fight global warming we need to develop effective methods to first identify timely and precisely heat stress in dairy cows and second to mitigate it both mechanically and or nutritionally historically heat stress research has focused on lactating cows and mitigating strategies to prevent heat stress to avoid heat stress focusing on early lactation and the breeding period to improve male production health and fertility outcomes but heat stress that not discriminate and it will take a toll on older cattle at all life stages including calves growing heifers dry cows which are not often considered for heat abatement on farm so we focus on the dry cow on the dry pregnant cow and why we do that is because during the dry period the mammary gland is undergoing two very important processes involution and redevelopment as they approach partition which are highly regulated processes and will set the stage for a successful next lactation so any disruption such as hyperthermia in this case heat stress can lead to a less productive other in the legs lactation this has been observed across 14 studies from our groups and others ranging from colgar in 1982 providing shape to their cows to more sophisticated active cooling techniques to promote evaporative cooling in our studies with fans and soakers but regardless of the heat abatement method which is denoted here in blue we can see that the average milk loss is approximately five kilos per day if these are not implemented now the tribute is important not only for mammary growth but it also coincides with the last trimester of gestation and this is when even though we know that all windows of development during gestation are important during this last trimester it's when 75 of the growth is occurring and is characterized by tissue accretion organ maturation and cell differentiation so this is the premise of fetal developmental programming which refers to how the intrauterine environment experienced by the developing fetus can shape the growth growth and physiology of this fetus and will determine phenotypes that will express in that offspring later in life so in the context of hyperthermia we can think of this as if you remember the fetus will have limited ability to regulate its own temperature while in utero so unfavorable conditions such as hyperthermia during this developmental window will derail always impact the organ development organ function physiological processes will alter behaviors and this can have long-term repercussions or consequences for future performance so those molecular signatures that are uh and physiological changes that are driven by in utero hyperthermia it's an active area in my lab of investigation in my lab today i'm going to focus on the phenotypic outcomes for the most part legislation heat stress induces reductions in gestation length on average we have seen four days and this is consistent across all our studies but one and we all always work with multiple scales here um calves is exposed to in euro hyperthermia are going to be born smaller on average 4.5 kilograms smaller and uh at weaning this growth disparity will remain and on average those uh born to heat stress dams are going to weigh seven kilos less and on those these the differences in body weight at first calvin of that heifer are similar these growth disparities are observed up to one year of age now um we also documented reductions in hip high width high body length head circumference so it's a little bit more than just body weight and more recently we reported differences in organ weights including immune related organs such as the spleen and the thymus those are smaller and metabolic and stress related tissues such as enlarged adrenal glands and also smaller mammary glands as even as a percentage of body weight so one of the most critical management factors as you might know in calf survival is feeding timely and enough good enough quality colostrum to that calf in order to improve positive strength so calves that are born to in utero hyperthermia have reduced apparent efficiency of immunoglobulin absorption from colostrum and this has been approximately between 10 to 20 percent and this leads to lower immunoglobulin circulation throughout the first weeks of life and as you can imagine this has a huge implication in their ability to develop and to mount a successful immune response but what caught my attention more even more when i was first uh studying this uh started studying these is this report by montero indicating that the daughters of late gestation heat stress stamps so when they were in neural heat stress they produced less milk in their first lactation so this is two years after they are born to a heat stress dam those first lactation heifers are going to produce less milk during um that first lactation across the first 35 weeks in milk so one of the first questions was that i have was whether this effect was in the offspring was transient was permanent do they recover what can we do about this so we recently published this study this manuscript in which we summarized 10 years worth of experiments at the university of florida where again working with multiparous cows under heat stress inside a barn or under active cooling with fans and soakers for the last 46 to 50 50 days of gestation so during that dry period so we were able to collect data from approximately 200 dams per group 80 daughters that we were able to follow for three lactations and approximately 20 grand daughters per group so these are granddaughters to the dam that experience heat stress during the gestation so we use happy farm records milky old protein fat and major life events we pull them together and we analyze them this graph shows the survival analysis of the daughters so we have the survival percentage here in the y-axis and h in days in the x-axis and here we have the different life events um we need first ai first calvin and first second and third lactation so it's clear to see that the red line which represents the in utero heat stress daughters it's always below the blue line which represents in your whole daughters so in summary um immutable heat stress daughters have reduced survival to first calvin have reduced survival through the third fourth and fifth year with our response to second first second and third lactation uh overall it has a once they enter the the cat for the first time and they enter the lactating string ascending heart the productive life is reduced by approximately five months and overall the lifespan of those cattle are reduced by 11 months if they are born to a heat stress dam now when we look at the milk production of those heifers that do make it to first uh the first lactation actually uh they we see that they make less milk on average 2.2 kilograms less so they start off producing less and across 35 weeks in milk we see again those uh impeding heat stress daughters producing this milk all along during those weeks and when we look at the second lactation we see the same pattern even though they start off the same they reach peak at different times and then the the red lactation curve it's always below the blue same pattern for the third lactation uh they produce less milk again they start off a little different here but then the red which are the neutral heat stress daughters they really crash after expectation so it's clear this data illustrates that neurohyperthermia which is experienced by the fetus can program long-lasting lower yields phenotypes with lifelong consequences on survival so it's a pretty big impact for a relatively short amount of time that they experience hyperthermia in euro now um the million dollar question or at least the one i'm uh searching for was trying to answer why this happened why this animal survived less why they produce this milk and so as a curious mammary physiologist i think the answer is in part in the marvel again at least the less productive the decrease in productivity so two years after we went in uh into those uh uh daughters when they started the first lactation and we collected multiple mammary biopsies at different stages of their lactation and what we found here is the euro cool mammary gland and this is in euro heat this is 21 at 21 days in milk so we found that the mammary gland of uh heat stroke in neural heat stress daughters um have less smaller alveoli as you can see here this is the alveoli so they were smaller when you compare them this is the same magnitude same days in milk so they had smaller alveoli with more connective tissue surrounding those alveoli lower proliferating cells percentage of proliferating cells and when we look closer at those alveoli we see this is the alveoli these are the mammary epithelial cells which are the cells producing secreting the milk we see that they have fewer those smaller alveolis were composed of fewer secretory cells so we believe this is one of the leading causes to reductions the reductions in male production that we observe of about 4.55 kilograms um so these alveoli have less secretory cells less um synthetic synthetic capacity and this is quite interesting now with this in mind we wanted to estimate the economic losses associated to those phenotypic outcomes and assess the repercussions to the theory the us their sector so we know that heat stress during lactation could cost 1.5 billion dollars in losses and this was estimated by some here in 2003 we know uh by ferreira in 2016 um that uh legislation heat stress in those dry cows um impact um an economic loss of 800 million losses um just by considering the reductions in milk heal up to down in the next lactation what we wanted to know is uh with the knowledge in hand that we had about the effects in the offspring how about those losses in the offspring in multiple lactations uh and how about their reductions in survival can we account for that somehow so assuming that no drag house the cows are not dry during the not cold during the dry period in the u.s and assuming those milk yield differences that we found in our studies in florida accounting for the number of heat stress days per state we calculated the annual losses associated with the daughters were born to dance that were heat strategies so this graph shows the 24 states with more dairy cows and florida which is the state with more heat stress days and we see the total million dollar loss per year per state and we separated by the cost associated with the extra heifer raring productive life and milk production so collectively when we sum up across all u.s we see that the economic loss of the offspring are estimated at 600 million dollars okay so this is pretty significant and the financial damage of the in euro offspring together with the financial loss of the dam can climb up to 1.4 billion dollars which is comparable to the estimations that we have for the loss in lactating cow so it's pretty significant yet this analysis does not include the toll that heat stress takes on legislation cows on the second generation of those so the granddaughters of those dams and also on the lippers heifers so all our studies have been done primarily focusing on multiple scales however um we wanted to look at this we wanted to look at oliver's heifers why are we doing this well heifers we always say they are still growing um you know they are diverting their energy towards their growth their field growth but their mammogram is growing exponentially at that time so we thought well maybe heat stress during that late gestation even though they don't have a dry period percent that narmigan is growing at an exponential rate so heat stress during that time could affect that impact production and also impact the pro progeny so can we call them how will they respond to these cooling methods that we use and so these are this is a picture of pregnant heifers in florida that are typically housed on pasture exposed to direct solar radiation and temperatures that exceed 35 degrees uh this is the experimental period from july to september and that leads to respiration rate that exceeds 100 bits per minutes during the hottest time of the day and vaginal temperatures that are well above 39.5 degrees celsius so um pretty intense so what we did is we moved those inside the barn for the last 60 days of the station where you can see that it's still hot but at least it's less than 30 degrees 35 degrees celsius just by protecting them from solar radiation so just the shade of the barn of the crystal barn here will decrease the respiration rate to 60 bits per minute and when we add active cooling uh with fans and soakers that respiration rate uh decreases even further to 45 bits per minute and we see uh the same trend in vaginal temperature although that during that hot hottest time of the day they don't uh pick as they do on posture so if we look at the sweating rate across a 36 hour period these heifers in red are really working a lot harder to dissipate heat even at night which is during this period yet they do it inefficiently because their skin temperature is still high their body temperature is still elevated relative to those heifers that we put them inside a barn and we provide object actively so clearly this will take a toll on them that is not transient they will produce less milk in the first lactation even if we cool them because this is the milk production the first 15 weeks in milk of that heifer and we see that if we don't cool them uh they lose 1.3 kilograms per day in milk they also have shorter gestation length on average three and a half days um just like multiple scales but we don't see any differences in birth weight in the body growth of those heifers uh during the pre-winning period but we do see impermanent immunity just like multiple scouts the apparent efficiency of absorption if these cats are born to uh in utero uh heat stress uh and also the immunoglobulins are going to be diminished across that pre-winning period so in summary legislation in heat stress in dry cows have multi-generational and multi-lational effects impacting the cow under heat stress their daughters and their granddaughters as well um nulliparous pregnant heifers feel the heat too after cooling them promotes milk yield and improves after an immune function um and while there's not much that we can do about the weather i mean at least in the short term there are things we can do to protect the animals from getting hot so what can we do what can be done to revert the long-term effects on the offspring on the progeny well we have um we had a very nice seminar from dr collier and dr cordoso showing the potential of mechanical cooling and nutritional strategies as well so we have shown that cooling works for dry cows it works for growing heifers now the question will be can we nutritionally program our offspring uh and our next speaker will be dylan talking about this um some in our next um part of this seminar but in general nutritional strategies have been mostly aimed at maintaining you know water balance nutrients electrolyte intake or to satisfy the needs of certain vitamins and mineral and we think that there's an unexplored opportunity here to supplement amino acids during pregnancy particularly those that serve as methyl donors such as methionine choline to promote maternal protein synthesis but also with the potential to program the offspring so with that my 20 minutes are um over so i thank you for watching this seminar and i will be open to questions after thank you thanks ximena for that excellent presentation and we certainly appreciate the opportunity to have you with us today you've given us many truly great insights to understand and use and i know you'll be ready to take some questions after the next presentation i just want to remind the audience to please post your questions so we have a good selection of questions to ask both speakers after the next presentation so next up is dr juan moore from the university of illinois who will share insights and research findings that help us further our understanding how of how nutrients impact heat stress one the floor is yours today we're gonna discuss the role of specific nutrients on potential benefits uh on the lactating cow during heat stress we're gonna try to address uh mechanisms um a little bit during this presentation we're also going to discuss potential avenues for um using these nutrients to uh help the cows not only do establish lactation but potentially during the transition period and also during pregnancy now let's just um revise very quickly some of the established knowledge in the context of metabolic and physiologic adaptations that occur in the cow during heat stress now feed intake decreases um during periods of heat stress that's been well known for uh quite a number of years uh but it hasn't been until uh recently uh when um uh we began to understand a little bit better some of the endocrine responses that uh after the metabolism of the cow during this period uh the most important one being the fact that insulin secretion increases substantially during heat stress which uh in turn prevents adipose tissue like policies and reduces the concentration of non-certified fatty acids that can be used as a source of energy for a number of tissues in fact this increase in insulin will essentially mean that the mammary gland has a lower availability of glucose which is going to also impact negatively milk production in turn muscle and other cells are going to make greater use of glucose to generate um a source that can allow them to synthesize atp uh so insulin action uh it's uh it's part or or or changes in insulin action are part of this new state that develops in the calder in heat stress uh muscle uh a characteristic uh tissue that responds to insulin is also going to be affected during heat stress uh in particular uh there's gonna be quite a bit of turnover of muscle protein which uh provides amino acids that can then be used by the liver for gluconeogenesis so again to try to meet the shortfall in glucose that is not coming from fermentation in the rumen but at the same time those amino acids are also going to be used for a number of other functions one of which is the immune response again and the liver being one of the important ones in that context some of those amino acids are going to be used to synthesize acute phase proteins which again is going to be part of of of the immune uh uh response of the of the animal during heat stress now our research of the last of the recent uh five to ten years has also uncovered that during heat stress uh the cow not only undergoes a period of reduced feed intake and outer endocrine and metabolic environment but also uh a potential uh period uh during which uh intestinal permeability could be affected now um if you think about it during heat stress one of the adaptations that the cow undergoes is uh besides the ones we discussed previously is it's the the attempt to release heat uh and in one of the methods whereby the cow attempts to do that is to uh channel blood flow away from the gut tissues uh to allow the animal to dissipate heat to the environment now that's obviously a survival mechanism that is important but uh in the context of gut gut physiology could potentially lead to a break decrease in the supply of nutrients for those cells to um to um to remain intact uh and in fact that's what's been seen in the in the last two years during heat stress there's an increase in intestinal permeability um partly because of the reduction of certain proteins that form complexes that allow those epithelial cells to remain together so to prevent or to form a barrier that would limit movement of luminal products for example toxins derived during the process of fermentation or absorption potential bacterial products such as lipopolysaccide um but again during heat stress uh because of the weakening of these junctions uh between cells there's a greater uh movement of these uh molecules into the circulation um and often they contribute to this uh inflammatory cascade that we uh mentioned in the previous slide now uh in addition to uh to the gut to the endocrine changes that um we know occur during heat stress we also need to keep in mind that individual organs and especially the metabolic uh important organs such as the liver the adipose and the mammary gland uh are also going to undergo undergo uh changes in the context of the immune response and also in the context of the antioxidant response now this is data that we generated uh in which we were trying to compare the response uh at the level of mrna abundance in cows that capped in the summer versus spring so this is data specifically for uh for liver i also want to draw your attention that in addition to the liver there's been some work published dealing with adipose tissue and even the mammary gland in the context of these molecular responses that can occur uh and that are associated with the immune system and the antioxidant response so each of these major organs is going to be responsive uh and in the case of the liver we know for a fact that there's a marked increase in the biomarkers that are associated with changes in the inflammatory response for example heptoglobin a classical acute phase protein serum amyloid uh and even lipopoly lipopolysaccharide binding protein these are important biomarkers of inflammation that are markedly upregulated during uh heat stress so again that will indicate that in fact this important organ is an immune response which again we need to take into account in the in the overall picture of this cow [Music] being fed during periods of heat stress in addition to these classical changes in inflammation and oxidative stress we also um uh uncovered that there are other important uh mechanisms that are potentially altered that are associated with some very specific nutrients for example we detected a large upregulation of metallotine that's an important enzyme that is involved in zinc copper and glutathione and homeostasis we're going to talk a little bit more about those in the next few slides we also saw a very large increase in the foliage receptor which of course would indicate that potentially during these periods of heat stress and also potentially associated with this inflammatory and oxidative stress response this nutrient might become very important in allowing these organs to adapt or to cope to these stressful situations in tissues like the adipose and the mammary gland again those tissues those cells are going to have uh they're going to undergo their own adaptations even at the molecular level and some of the data we have indicate that there's an antioxidant response taking place and again that means that potentially nutrients or the nutrients applied it's also going to be important in the context of allowing those pathways to to help those cells respond to this heat stress now so in the context of heat stress really we need to think about two major aspects of the physiology of the animal or three if you include the immune system so we need to think about the metabolic response we need to think about the immune response and we need to think about the antioxidant response now there's been quite a bit of work um to uh in in the context of trying to alleviate some of the changes that we often see during heat stress related to energy violence uh specifically the the the function of insulin in during this uh period of heat stress and in that context the use of chromium it's uh has received quite a bit of attention historically chromium is one of those uh micronutrients that has been well known to have a role in the context of insulin signaling okay so there's a lot of aiding monogastrics and even humans uh supporting the supplementation of chromium to uh enhance uh insulin function we don't necessarily know the mechanisms very well but between some working dairy cows during heat stress using chromium here's one um data set from an experiment that actually looked at two levels of chromium relative to unsupplemented control cows again it was done during moderate um heat stress conditions in the summer and as you can see here from this data clearly the medium dose of chromium methionine had a beneficial effect not only in the context of energy corrected milk but also in the context of milk fat and protein concentration and even feed intake so these data are important because again a big driver of energy balance is obviously feed intake and uh in in the context of heat stress so clearly some beneficial effects associated with chromium the data that they reported on nifa and insuli also potentially suggested that in fact this level of chromium supplementation actually had a beneficial effect in the context of helping re-establish if you will uh proper insulin action in these animals so nephi were lower uh even when feed intake was higher and insulin was reduced in terms of plasma concentration so again they didn't report uh much in terms of mechanisms but these data uh again uh are somewhat similar to what you would see in monogastric animals in the context of what insulin how insulin function might respond to chromium now these data were replicated in another experiment that use a much larger number of animals so there's some good evidence for the role of additional chromium during heat stress in the context of uh better insulin function now let's move and talk a little bit about the the other uh major component of the adaptations of the cow during heat stress and this has to do with with uh with immune function and also antioxidant function those two are related now let's talk a little bit about uh what we know about the typical antioxidants uh that are commonly fed to dairy cattle and here you have a list obviously some of the water-soluble vitamins are well known to be um to be potent antioxidants there are some trace minerals that we know are also important in the context of an antioxidant mechanism either because they are co-factors for enzymes for example selenium and glutathione peroxidase manganese with superoxide dismutase so some of these antioxidants either work as co-factors for potent enzymes that help in the antioxidant response or they can actually scavenge uh radical um species such as the case of vitamin c so again there's a there's uh there's a a good body of knowledge in the context of these different uh antioxidants now one of the um important aspects of the antioxidant mechanisms is that uh cells um and tissues and um like the liver for example are capable of also synthesizing uh potent antioxidants and endogenously and i'm referring particularly to taurine and glutathione here as part of the transformation pathway uh some of the work that's been done um with the room and protected methionine to increase the post-romanian supply of methionine has consistently shown an increase in the concentrations of glutathione and often the concentration of taurine also not only in liver tissue but also in the circulation and that agrees with the the known uh pathways whereby methionine and other metal donors like choline and betaine can potentially um allow the animal to generate some of these but important antioxidants now i also want to highlight that in addition to methionine and some of these other methyl donors as uh serving as substrates as we can see from the from these uh pathways uh there are other uh micronutrients that can also be required in order to run these pathways in a more optimal fashion particularly during heat stress that's the case of zinc and copper which happened to be cofactors for a couple of important enzymes folic acid also feeds into this into the skin and of course the water-soluble vitamins which we know are are also part of some of these enzyme complexes so it becomes a little bit more complex to think about these endogenous synthesis of antioxidants but these are mechanisms that the animal also can use during stressful periods now let's go through some examples of what's been reported in the context of use of specific nutrients there's been an interest in zinc for obvious reasons zinc is an important nutrient for the immune system zinc also can potentially trigger some molecular adaptations because there are some zinc sensitive transcription factors if we can call them that uh in this particular experiment that was done on their fairly fairly moderate to severe heat stress even though there was no difference in the context of dry matter intake on energy corrected milk or other performance measurements during heat stress uh the authors were able to detect an increase or a tendency really for increase in this important tight junction protein referred to as ekadiri remember when we were talking about the features related to um uh intestinal permeability we we mentioned the fact that uh cells of the epithelium uh in in the gut are held together through uh junction proteins that allow them to to again remain in place and ecavirin happens to be one of those junction proteins in this particular example um rna mrna abundance of these protein in the memory samples that they were able to harvest really showed a beneficial effect of this zinc methionine complex that replaced a portion of the zinc hydroxychloride supplement that was being fed so again potentially some of these nutrients may not cause changes in the performance during heat stress but potentially they can have beneficial effects at uh at specific uh sites in in the animal be it the intestine be in the liver be it the adipose or be it the mammary gland uh selenium has is is another important nutrient that has been evaluated in the context of heat stress of course selenium as we saw from the previous slides is an important component of the antioxidant systems in this particular experiment these authors actually use a selenium methionine complex in place of selenium selenide which uh it's it's a common form that's fed to dairy cows to induce heat stress they actually put cows through a hay chamber and they were able to achieve a fairly severe level of heat stress during a period of nine days so some of the data that they were able to measure during this period relate to the activity of glutathione peroxidase which we can see here uh remained fairly stable and was even greater for cows in the celine methionine product towards the end of that heat heat stress period now in some experiments done in monogastrics it's it's been demonstrated that selenium can actually increase not only the abundance of glutathione peroxise but also its activity and this data seem to indicate that mechanisms similar to the ones that have been published with other species also seem to work here in the cow uh in the context of of the concentration of circulating lipid peroxidation products we have a couple of data points here that indicate that selenium methionine did have a beneficial effect one of them was the substantial decrease in hydrogen peroxide uh which again would indicate that in fact these antioxidant systems due to feeding selenium methionine were activated or working in a more uh optimal fashion now there's another um lipo peroxidation product that can be measured uh through the mda assay and this is what we have here also in the in in the graph and again as you can see uh cows that were fed the selenium methionine product actually had a gradual decrease in concentrations of these mda molecules that became significant and reach on a deer uh towards the end of that heat stretch period so clearly selenium methionine in this particular experiment was um more beneficial to the animal in the context of helping alleviate alleviate the antioxidant uh or the ox the the oxidative stress uh response in the animals during heat stress and in fact we need to point out that some of that um benefit appears to have been because the selenium in the selenium methionine complex that was fed seem to be more bioavailable and we can gauge that because milk total selenium was higher in calcite the selenium methionine complex so clearly data um from this experiment suggests a potentially important role of selenium in the context of the antioxidant response and if in fact the antioxidant response increases during heat stress that means that immune response or the inflammatory cascade is also going to be diminished now moving uh to some of the data that we have generated in the context of methyl donors and the immune system this is a table from a review paper that we published uh last year and again just to give you an idea of what we summarize in this table uh it was to emphasize uh some of the responses that we have detected and other groups have detected working with uh sources of ruin protective methionine in the context of biomarkers of oxidative stress and inflammation and again you can see the table here and as i indicated before consistently we've seen that cows that are fed room and protected methionine or hydroxy analog are actually able to synthesize more glutathione and we've been able to detect that through measurements of liver tissue and also plasma so they're able to to channel some of the methionine uh through that antioxidant antioxidant pathway that i indicated before in addition to again being able to to use more of that methionine for milk protein synthesis which we have also demonstrated occurs when when the supply of methane increases uh in this particular fashion so again we've seen some clear benefits in the context of transition cows and um and the potential antioxidant response during those stressful conditions in the animal there's been less work looking at methyl donors during heat stress but this is some data that was published uh fairly recently in which authors were trying to use a uh or induce and a heat stress uh challenge through the um the use of heat blankets uh on cows and here i have uh i'm showing some some data that uh to allow you to see some of the main uh uh results that they were able to uh detect now the baseline period in that baseline period or all cows were in fear more neutral conditions and you can see here that um protein percent was clearly uh higher in the calcium and protected methionine but other aspects of performance and uh and and were not really affected uh statistically at least by feeding roman protective methionine now the data that i'm showing here during during the test period uh pertains to the response relative to that baseline period so for example during heat stress cows that would that were fed room protected methane hara not only were able to sustain uh higher concentrations of fat but those concentrations were statistically higher in those animals relative to the pair fed um animals now pear feeding is a it's it's a common way in which we can try to uh remove uh the the potential effect of our reduction in feed intake during heat stress so we typically can can can have perfect animals and compare those to the heat stress animals so you perfect to the level of intake that heat stress cows go to and then again you're able to remove that from from your interpretation so again clearly ruling protected mentality during heat stress had a beneficial effect in the context of five percent and even in the context of casein percentage uh again this study did not report any specific mechanisms but uh what we can say with confidence is that uh the positive effect that was detected for roman protected methionine at least at the performance level was likely related to to the to the to the fact that those cows uh had an uh adequate supply of methionine in the mp um we have tried to address the issue of methane of methanol and calling supply to metal important metal donors during heat stress by using an in vitro approach and uh in we've done a couple of cell types these these are data from immune cells uh in the form of neutrophils and what we try to do here was to look at different levels of choline and different ratios of lysine to methionine in vitro in the context of the heat stress response and the way in which we induce heat stress was we had these cells that were isolated from mid-lactation mid-lactation cows would put them through a two-hour heat stretch period during which they were they received these different these different treatments now what i'm showing here is just to emphasize a couple of things first that that heat stress does affect the functionality of the neutrophils and we can see that because uh when we measure the il-6 concentration there was a decrease uh from the thermal neutral uh conditions to the heat stress in the context of of these um of these inflammatory cytokines so lower concentration told us that again heat stress did have a negative impact now the other reason why we can see that this heat stress did have an effect is when we look at the expression of heat shock protein 70 a classical biomarker of heat stress it was actually quite quite high um during heat stress the important thing is to realize that when we increase the supply of choline in particular we were able to detect an increase in heat shock protein 70. now on increasing this in this uh protein it's actually a beneficial response that the cell is is going through to try to restore um homeostasis uh in in in the cells uh that again are potentially is potentially altered due to the heat stress so we take this response to mean that choline in particular but potentially also methionine could have a potentially important impact during heat stress at the level of immune cells and that can be uh somewhat supported by the fact that for example for methionine the concentration of il-6 was actually increased to above those initial concentrations uh during heat stress when we increase the supply of this uh amino acid uh in in the in the culture medium so the question is now are these data also relevant to the potential to what may happen in vivo that's something we haven't tested yet but uh some of these uh data that we generated seem to indicate that in fact methionine will have a potentially important role along with choline in the context of immune cell function uh with that i'm going to leave you with some important uh key take-home messages from my perspective uh for you to keep in mind in the context of how can how potentially important certain nutrients can be during heat stress we need to think not only about a single type of nutrient or nutrient uh but we need to think about the fact that because we have different cell systems that are important um in the animal and because we know that different cell systems have different uh enzyme and molecular machineries uh potentially we need to we need to we're gonna have to think about uh not only a single nutrient but a mixture of nutrients and being able to deliver uh the correct amounts or the most beneficial amounts during heat stress so again that means that this is a complex question remember that when we're thinking about heat stress and potentially alleviating problems associated with this stress in the animal through nutrition we need to think about metabolism and a number of non-metabolic aspects including the antioxidant response and also the the inflammatory response and remember that when we have uh it could even be more complicated because we may have uh cows that are pregnant we may have cows that are um are giving birth during the transition period so that those are additional stressors and we're talking about pregnant animals um we're we're also talking about potentially diminishing the supply uh to the fetus or to the growing embryo and that's that's also something that we need to we're going to have to address especially when we're thinking about animals that are calving around that are calving around summertime or during heat stress conditions uh it's important that we not only think about dietary supply of of nutrients that are cancerous antioxidants but remember that we can trigger the the or increase the synthesis of endogenous antioxidants glutathione and taurine in particular uh and especially through the uh higher supply or increasing the supply of methyl donors uh clearly we we're just scratching the surface in the context of the specific nutrient effects uh in the context of heat stress this is this is a this is an area that's fertile that we are going to be generating a lot more knowledge in the near future and i will leave you with um some thoughts in the context of feed additives that are starting to show some potentially important beneficial effects in the context of heat stress and those are the ones that are derived primarily from yeast or mixtures of microbial uh products and vitamins some evidence in the literature that potentially those can have a beneficial effect at the level of of the cow during heat stress and again we we need to think about those potentially in combination with some of these other nutrients when we're trying to design diets that can help cows um cope with heat stress during during uh during the summer and with that i thank you for your attention