CN108719194B - Method for enhancing antioxidant and immune functions of lambs - Google Patents
Method for enhancing antioxidant and immune functions of lambs Download PDFInfo
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- CN108719194B CN108719194B CN201810581999.1A CN201810581999A CN108719194B CN 108719194 B CN108719194 B CN 108719194B CN 201810581999 A CN201810581999 A CN 201810581999A CN 108719194 B CN108719194 B CN 108719194B
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/02—Breeding vertebrates
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/174—Vitamins
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/10—Feeding-stuffs specially adapted for particular animals for ruminants
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Polymers & Plastics (AREA)
- Zoology (AREA)
- Animal Husbandry (AREA)
- Environmental Sciences (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Animal Behavior & Ethology (AREA)
- Biodiversity & Conservation Biology (AREA)
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Abstract
The invention discloses a product and a method for enhancing the antioxidant and immune functions of lambs by adding nicotinamide to dairy goats in the perinatal period and application thereof, and belongs to the technical field of nutritional feed additives for dairy animals. Specifically, the nicotinamide is added to the dairy goat in the perinatal period for feeding, so that the addition of the nicotinamide has no influence on the birth weight of the lambs and the growth and development of the 28-day-old lambs, and the gestation period of the ewes; but obviously improves the oxidation resistance and the immunity of the lambs and lays a foundation for exploring the regulation and control effect of the B vitamins and nicotinamide nutrition of a parent on offspring.
Description
Technical Field
The invention belongs to the technical field of nutritional feed additives for dairy animals, and particularly relates to a product and a method for enhancing the antioxidant and immune functions of lambs by adding nicotinamide and/or nicotinic acid to dairy goats in a perinatal period and application of the product and the method.
Background
Perinatal is one of the most critical physiological periods of dairy animals, which goes through the transition from pregnancy to parturition and then to lactation, and has large changes in physiology, nutrition, metabolism and immunity, and frequent metabolic and infectious diseases. In addition, the dairy animals can have appetite suppression in the later gestation period, and the dry matter feed intake after delivery can not meet the nutritional requirement. Energy intake of dairy animals is reduced due to the reduction of dry matter intake, and energy demand is increased due to embryo growth and lactation, resulting in negative balance of nutrients including negative energy balance, negative protein balance and negative vitamin balance during perinatal period. The milking animals can ensure the milk yield in the lactation period by regulating the metabolism of the organism. At present, a plurality of achievements exist in the regulation and control research aiming at the negative balance of nutrients in dairy animals, particularly in the perinatal period, and the functions of part of nutritional feed additives (such as rumen bypass choline and rumen bypass methionine) are proved, so that the negative balance of energy and protein can be effectively relieved, and the health and the lactation performance of animals are improved.
The overnutrition, the insufficient nutrition or the unbalance of the maternal in the gestation period and the lactation period can change the organ functions of the offspring and reduce the production performance of the offspring. The maternal effect refers to the phenomenon that the physiological status and production performance of the offspring livestock and poultry are directly affected by the female parent. With the development and deep research of animal nutrition, genetic breeding and reproduction, and bioinformatics, the research finds that the maternal effect is not only related to genetic information, but also closely related to environment and nutrition level. Since fetal genetic programming determines the structure, function and metabolism of the body, nutritional deficiencies during pregnancy are a source of post-adult metabolic disease development. Maternal nutrition plays a crucial role in fetal growth and is also the basis for normal growth, development and reproduction of the animal after adulthood. Researchers have conducted extensive experimentation to determine the nutritional needs of animals, but the optimal nutritional supply during pregnancy remains a significant problem for many species in the world (e.g., swine, cattle and sheep, etc.).
Nicotinic acid and niacinamide both belong to the B vitamins, collectively referred to as vitamin PP. Nicotinic acid, also known as nicotinic acid, formula C6H5NO2(ii) a Nicotinamide, also known as nicotinamide, of formula C6H6N2And O. It is believed that nicotinic acid is converted into nicotinamide in liver or other tissues, and the nicotinamide and nicotinic acid have common vitamin activity, can participate in the regulation and control process of body energy metabolism, and has antioxidant and anti-inflammatory effects. Research shows that the nicotinamide can relieve negative energy balance of dairy animals and improve animal health, but no research report exists so far on whether growth and development, oxidation resistance, immune function and the like of offspring can be influenced.
Disclosure of Invention
Aiming at the defects of the prior art, the inventor provides a product and a method for enhancing the antioxidant and immune functions of lambs by adding nicotinamide to dairy goats in perinatal period through long-term technical and practical exploration, and application thereof. Specifically, the nicotinamide is added to the dairy goat in the perinatal period for feeding, so that the pregnancy period of the ewe, the birth weight of the lamb and the growth and development of the 28-day-old goat are not affected, the antioxidant capacity and the immunity of the lamb are obviously improved, and a foundation is laid for exploring the regulation and control effect of the B vitamins and the nicotinamide nutrition of the maternal goat on the offspring.
The invention aims to provide an application of nicotinamide/nicotinic acid added to a perinatal milk goat to enhance the antioxidant and immune functions of the lamb.
The invention also aims to provide a nutritional feed additive for enhancing the antioxidant and immune functions of lambs by adding the nutritional feed additive to the dairy goats in the perinatal period.
The invention also aims to provide a using method of the nutritional feed additive.
In order to achieve the purpose, the invention adopts the following technical scheme:
in a first aspect of the invention, the application of nicotinamide/nicotinic acid added to a dairy goat in a perinatal period to enhance the antioxidant and immune functions of the lamb is provided;
wherein the perinatal period comprises the later gestation period and the early lactation period of the dairy goat;
further, the perinatal period is from 21 days before delivery to 28 days after delivery of the milk goat;
the addition amount of the nicotinamide and/or the nicotinic acid is 3-8 g (preferably 5 g) of nicotinamide and/or nicotinic acid fed to each perinatal milk goat every day;
preferably, the adding method is carried out in a direct drenching mode, so that the intake of nicotinamide (nicotinic acid) is ensured.
In a second aspect of the present invention, there is provided a nutritional feed additive for enhancing antioxidant and immune functions of lambs by adding to perinatal dairy goats, the nutritional feed additive comprising nicotinamide and/or nicotinic acid;
preferably, the perinatal period comprises the later gestation period and the early lactation period of the dairy goat;
further, the perinatal period is from 21 days before delivery to 28 days after delivery of the milk goat;
in a third aspect of the present invention, there is provided a method of using the nutritional feed additive described above, the method of use comprising: feeding the perinatal milk goat with nicotinamide/nicotinic acid daily;
preferably, the feeding method is carried out in a direct drenching mode, so that the intake of nicotinamide (nicotinic acid) is ensured;
preferably, the feeding amount of the nicotinamide/nicotinic acid is controlled by feeding 3-8 g (preferably 5 g) of the nicotinamide/nicotinic acid to each perinatal milk goat every day.
Compared with the prior art, the technical scheme of the invention has the following beneficial effects:
(1) the invention is researched for the first time and finds that the nicotinamide is added to the dairy goat in the perinatal period, so that the nicotinamide has no influence on the birth weight of the lambs and the growth and development of the 28-day-old goats, and has no influence on the gestational period of the ewes; but obviously improves the oxidation resistance and the immunity of the lambs, fully indicates the complexity of a nicotinamide regulation mechanism, and simultaneously lays a foundation for exploring the regulation and control effect of B vitamins nicotinamide nutrition on offspring;
(2) the research of the invention shows that the nicotinamide added to the ewe in the full period of perinatal period reduces the transmission of active oxygen free radicals to the fetus through the placenta. The nicotinamide is added to the ewes in the late perinatal period and the whole period, so that the oxidation resistance of milk is improved, active oxygen free radicals are reduced and are transmitted to offspring through the milk, the oxidative stress of lambs is reduced, and the oxidation resistance of the lambs is improved;
(3) the research of the invention shows that the addition of nicotinamide to ewes in the late perinatal period and the full period improves the immune function of lambs, and the improvement of the immune function is possibly related to the methylation level change of DNA;
in conclusion, the invention combines the negative energy balance problem of the ruminant in the perinatal period with the maternal nutritional effect, researches the influence of the variation of the nutritional level of the dairy animals in the perinatal period on the oxidation resistance and the immune function of offspring, specifically researches the regulation and control effect of B vitamins and nicotinamide on the offspring, lays a theoretical foundation for further researching the regulation and control effect of nutritional feed additives such as vitamins and the like on the offspring, and has good application value.
Drawings
FIG. 1 is a graph showing the effect of nicotinamide addition to a dairy goat in perinatal period on the relative expression level of placenta antioxidant gene mRNA;
FIG. 2 is a graph of the effect of nicotinamide addition by perinatal milk goats on the relative expression of mRNA of placental SIRT 1;
FIG. 3 is a graph showing the effect of nicotinamide addition to dairy goats in perinatal period on the relative expression level of lamb liver antioxidant gene mRNA;
FIG. 4 is a graph showing the effect of nicotinamide addition to a perinatal milk goat on the expression level of inflammatory cytokines in the 28-day-old spleen of a lamb;
FIG. 5 is a graph of the effect of nicotinamide addition to a perinatal milk goat on spleen acquired immunocytokines in a 28-day-old lamb;
FIG. 6 is a graph showing the effect of nicotinamide addition to a perinatal milk goat on the lymphoinflammatory cytokine expression level of a 28-day-old lamb;
FIG. 7 is a graph of the effect of nicotinamide addition to perinatal milk goats on lymphoacquired immune cytokines in 28-day-old lambs;
FIG. 8 is a graph showing the effect of nicotinamide addition to perinatal milk goats on the expression of spleen DNMT1 mRNA.
Detailed Description
It should be noted that the following detailed description is exemplary and is intended to provide further explanation of the disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.
The present invention is further illustrated by the following specific examples, which are provided for the purpose of illustration only and are not intended to be limiting. If the experimental conditions not specified in the examples are specified, they are generally according to the conventional conditions, or according to the conditions recommended by the sales companies; the present invention is not particularly limited, and may be commercially available.
Examples
1 materials and methods
1.1 design of the experiment
In the test, 15 gestational milk goats of 2 fetuses are selected, the gestational milk goats are divided into 3 groups according to weight and feed intake matching, the control group (C without nicotinamide addition), the perinatal late group (P with nicotinamide addition from 1 to 28 days after delivery) and the perinatal full-period group (EP with nicotinamide addition from 21 d to 28 d after delivery) are respectively used, and 5 g of nicotinamide is fed at a ratio of 7:00 am every day in a drenching mode. After the lambs are born, 5 lambs are respectively selected from each group, and the corresponding groups are respectively LCGroup, LPGroup sum LEPAnd (4) feeding the breast milk of the corresponding treatment group by adopting manual quantitative feeding.
1.2 Breeding management
Animal experiments are carried out in a stock teaching test base of northwest agriculture and forestry science and technology university, and breeding is carried out in a colony house in a scattered mode. The basal ration was fed as a Total Mixed Ration (TMR), three times a day, 8: 00. 14:00 and 18:00, and freely eating and drinking water to ensure that the daily ration remains. Milking the ewe 2 times every day after calving (8: 30, 17: 30), and feeding the ewe with prenatal daily ration and postnatal daily ration according to different stages of a sheep farm feeding system.
After the fetus comes out, mucus on the body, especially mouth and nose, is removed in time, asphyxia death is prevented, information recording such as weight, sex, birth date and the like is carried out, and enough colostrum is fed within half an hour. The method adopts a separate milk basin to manually feed quantitative breast milk: when the ewes express the milk, collecting the breast milk according to groups; the breast milk is mixed evenly according to groups, heated to 74 ℃, then cooled to 37 ℃ and fed to lambs. Four times daily feeding, 5:00, 10:00, 15:00, 20: 00. The feeding amount is increased according to the body weight in a proper amount every week, and the feeding amount is slightly more in the morning and evening.
1.3 blood sample Collection
Jugular vein blood sampling is carried out at 14 days and 28 days of age respectively and 3 hours after morning feeding. Collecting blood with 5 ml vacuum blood collecting tube containing anticoagulant (heparin sodium) and anticoagulant-free blood collecting tube, storing in ice box at low temperature, rapidly taking back to laboratory, centrifuging, and packaging. And (3) carrying out water bath on the vacuum blood collection tube added with the anticoagulant at 37 ℃ for 10 min, placing the vacuum blood collection tube in an inclined mode until serum is separated out, centrifuging the blood collection tube at 3000 r/min for 15 min, collecting the serum and subpackaging the serum in a 2 ml centrifugal tube, and storing the serum at-80 ℃ to be detected. Centrifuging the vacuum blood collection tube without anticoagulant at 3000 r/min for 15 min, collecting and subpackaging the plasma in a 2 ml centrifuge tube, and storing in a refrigerator at-80 ℃ to be tested.
1.4 milk sample Collection
Milk samples were collected on the day of parturition, 7 days postpartum, 14 days, 21 days, and 28 days, respectively. Milking 2 times daily (8: 30, 17: 30), mixing twice a day and night milks in proportion, and subpackaging in 4 ml, 10 ml and 50 ml test tubes, and storing in a refrigerator at-20 deg.C for testing.
1.5 tissue sample Collection
After delivery of the ewe, the placenta and the placenta are generally discharged within 1 to 2 hours, the head of the ewe is fixed after delivery, scissors, a porcelain plate, tweezers, gauze, normal saline and the like are prepared in advance, the placenta is placed on the porcelain plate after being discharged, the placenta is cleaned, a proper amount of tissues are cut, and the placenta is placed into a 2 ml cryopreservation tube and a liquid nitrogen tank.
Lambs were slaughtered at 28 days of age and sacrificed by cervical bleeds. After slaughter, spleen, liver, lymph node and other tissues are taken down from the animal, the connective tissue, fat and blood clots on the surface of the animal are firstly stripped, then the animal is cleaned for 2 times by physiological saline, wiped clean by absorbent paper, placed into a culture dish, cut into pieces by scissors, collected into 2 ml freezing tubes and placed into a liquid nitrogen tank, and all the operation steps are carried out on ice. After the tissue sample is brought back to the laboratory at-80℃And (5) storing the sample to be tested in a refrigerator.
1.6 index determination
1.6.1 growth Performance
The initial weight of the lambs and the weights of the lambs at 1, 2, 3 and 4 weeks before morning feeding were measured. The time of lambing of the ewes was recorded and the number of gestational days was calculated. Lambs were sacrificed at 28 days of age using cervical exsanguination. Body weights were weighed before slaughter to allow calculation of organ indices. The spleen and liver tissues were removed from the animals, and the connective tissue, fat and blood clots on the surface were first stripped off, then washed 2 times with physiological saline, wiped clean with absorbent paper, and then weighed.
1.6.2 plasma and milk sample indices determination
The nicotinamide content in the plasma was determined by means of a high performance liquid chromatograph (Breeze 22695, usa) from Waters, using a fluorescence detector with an excitation wavelength of 322 um and a detection wavelength of 380 um; the chromatographic column adopts a C18 chromatographic column, and the parameters are 5 mu m and 4.6 multiplied by 250 mm; the amount of sample was 20. mu.l, and the flow rate was 0.8 ml/min.
The Growth Hormone (GH) content in plasma was determined by an enzyme-linked immunosorbent assay using a kit of Beijing Huaying institute of biotechnology (batch No. 20171022).
The kit for measuring the total antioxidant capacity (T-AOC) of blood plasma and measuring glutathione peroxidase (GSH-PX), superoxide dismutase (SOD), Catalase (CAT) and Malondialdehyde (MDA) is purchased from Nanjing Biotechnology engineering Co., Ltd (the product numbers are respectively A001-3, WST-1, A005, A001-3, WST-1 and A007-1, and the visible light method and A003-1), and the specific measuring process refers to the kit specification.
Measurement of immunoglobulin IgG in blood was carried out by ELISA using the Goat ImmunoglobLin G (IgG) Elisa Kit purchased from Nanjing Tanke Ltd. The kit adopts a competition method to detect the content of immunoglobulin G (IgG) in a sample. The measurement of interleukin 6 (IL-6) and TNF-alpha was carried out using an ELISA kit. The kit adopts a double-antibody one-step sandwich enzyme-linked immunosorbent assay (ELISA).
Total Protein (TP) was measured using a BCA protein quantification kit (cat. No.: A405-4, microplate assay). According to the serum (slurry): physiological saline =1: and 49, diluting to be tested. Tissue samples by weight (g): adding 9 times volume of physiological saline according to the volume (ml) =1:9, mechanically homogenizing in an ice-water bath at 3000 r/min, centrifuging for 10 min, taking supernatant, diluting with physiological saline at 1:9 to obtain 1% tissue homogenate, and testing.
Before the milk sample index is measured, the sample is pretreated. Collecting milk sample from-20°C taking out the refrigerator, melting, and then 4°C. Centrifuging for 30 min at 2000 g, removing upper fat, retaining whey, and packaging into 1.5 ml centrifuge tube. The antioxidant index in milk is the same as the antioxidant index in blood sample;
the measurement of IgG and cytokines IL-6 and IL-1. beta. in colostrum and normal milk is consistent with the measurement of the same indices in blood as described above.
1.6.3 relative quantification of tissue-like antioxidant genes
Total RNA of placenta and liver tissue was extracted using Trizol kit (purchased from Takara corporation), and the procedure was as follows with reference to the instruction:
(1) taking about 0.1 g of tissue sample, putting the tissue sample into a 2 ml centrifuge tube without RNase, adding 1 ml of Trizol, homogenizing by an electric homogenizer for 60 s, and standing at room temperature for 15 min;
(2) correspondingly adding 0.2 ml of chloroform into 1 ml of Trizal, tightly covering a bottle cap, fully performing vortex oscillation, and standing for 5 min;
（3）12000 r/min，4 °c, centrifuging for 15 min, and taking 400-500 mu L of supernatant into a new RNA enzyme-free EP tube;
(4) adding isopropanol (400-;
（5）12000 g，4 °centrifuging for 10 min, discarding the supernatant, and keeping the precipitate;
(6) adding 500 μ L of precooled 75% ethanol to wash the precipitate, shaking to make the precipitate float, and standing for 90 s;
（7）7500 g，4 °c, centrifuging for 5 min, and removing supernatant;
(8) repeating the steps (6) and (7), reversely buckling the EP pipe on the absorbent paper, and drying at room temperature for 5-10 min;
(9) adding 50-100 mu of LDEPC water to dissolve RNA;
（10）-80 °and C, storing to be tested.
The concentration of RNA was determined by means of a spectrophotometer (Nanodrop ND-2000) and the OD was noted260/OD280And OD260/OD230(ii) a Performing RNA reverse transcription, and detecting the relative expression of the tissue-like antioxidant genes; all primers used in the test are synthesized by Xian engine biosynthesis company Limited, and the parameter information is shown in Table 1.
TABLE 1 target genes and reference primers
Reverse transcription of RNA Using PrimenScriptTMRT reagent kit (purchased from Takara) was performed under 37 reaction conditions°C，15 min；85 °C，5 s；4 °C，Forever；
The Real-time PCR adopts SYBR Green chimeric fluorescence detection method and SYBR Premix ExTaqTMII kit (from Takara). The reaction conditions are as follows: pre-denaturation 95°C, 30 s; PCR reaction 95°C，10 s，60 °C，30 s，72°C, 30 s (40 cycles); dissolution curve 72°C, 5 min, three parallel wells per sample.
1.6.4 determination of expression quantity of tissue-like immune-related factor
The mRNA level expression levels of immunocytokines (IL-1 beta, IL-6, IL-8, TNF-alpha) and DNA methyltransferases in spleen, lymph and intestinal mucosa were measured by Real-time PCR. All primers used in the test are synthesized by Xian engine biosynthesis Co., Ltd, and the parameter information is shown in Table 2.
TABLE 2 target genes and reference primers
1.7 statistics of data
The experimental data were initially statistically organized using Microsoft Excel 2010, followed by data analysis using IBM SPSS 23.0, using One-way ANOVA, and multiple comparisons using Least Significant Difference (LSD).
2 results
2.1 Effect of Nicotinamide addition to Dairy goats in perinatal period on growth Performance of lambs
2.1.1 initial weight and body weight
From Table 3, with LCGroup comparison, LPGroup sum LEPThe group has no significant influence on the birth weight of the newborn lambs, the body weights of 1, 2, 3 and 4 weeks old and the average daily gain weight (the weight gain of the newborn lambs is not increased by the group: (P>0.05）。
TABLE 3 Effect of peripartum Nicotinamide addition on lamb birth weight and body weight
1LC= control, LP= group around postpartum period, LEP= perinatal full period group.
2.1.2 organ index
From Table 4, with LCGroup comparison, LPGroup sum LEPNo significant effect on liver and spleen organ index after 28-day-old lamb slaughter (P>0.05）。
TABLE 4 Effect of Nicotinamide addition on lamb organ index in perinatal period
1LC= control, LP= group around postpartum period, LEP= perinatal full period group.
2.1.3 Ketone body weight
From Table 5, with LCGroup comparison, LPGroup sum LEPHas no significant influence on the carcass weight of the slaughtered 28-day-old lambs (P>0.05）。
TABLE 5 Effect of perinatal Nicotinamide supplementation on lamb 28 day-old Ketone body weight
1LC= control, LP= group around postpartum period, LEP= perinatal full period group.
2.1.4 plasma growth hormone content
From Table 6, with LCGroup comparison, LPGroup sum LEPPlasma growth hormone content of 14-day-old and 28-day-old lambs in groupThe amount has no significant influence (P>0.05）。
TABLE 6 Effect of Nicotinamide addition on growth hormone in lamb plasma in perinatal period
1LC= control, LP= group around postpartum period, LEP= perinatal full period group.
2.1.5 Nicotinamide content in blood
From Table 7, with LCGroup comparison, LPGroup sum LEPThe group tends to increase the nicotinamide content in the serum of the 14-day lambs (P=0.092）。
TABLE 7 Effect of Nicotinamide addition to ewes in perinatal period on Nicotinamide content in serum of 14-day-old lambs
1LC= control, LP= group around postpartum period, LEP= complete period of perinatal group
2.1.6 gestation period
From Table 8, with LCGroup comparison, LPGroup sum LEP(the group has no significant influence on the gestation time of the ewes: (P>0.05）。
TABLE 8 influence of Nicotinamide addition in perinatal period on gestation time of ewes
1LC= control, LP= group around postpartum period, LEP= perinatal full period group.
2.2 Effect of Nicotinamide addition to Dairy goats in perinatal period on lamb antioxidant capacity
2.2.1 placenta Oxidation resistance index
From Table 9, with LCGroup comparison, LEPGroup significantly improved placental T-AOC: (P<0.05), no significant effect on the enzymatic activities of SOD and CAT: (P>0.05）。
TABLE 9 influence of Nicotinamide addition on antioxidant index of placenta in perinatal period
1LC= control, LEP= perinatal full period group. The difference of the lower case letters on the same row of shoulder marks is significant (P<0.05）。
2.2.2 placenta antioxidant enzyme mRNA relative expression level
As can be seen from FIG. 1, and LCGroup comparison, LEPGroup significantly reduced the mRNA expression level of placental GPX1 (P<0.05), has no significant effect on the expression of CAT, SOD1, SOD2 and GPX3 (P>0.05）。
2.2.3 relative expression level of mRNA of placental SIRT1
As can be seen from FIG. 2, and LCGroup comparison, LEPNo significant effect on the relative mRNA expression level of placental SIRT1 (P>0.05）。
2.2.4 Oxidation resistance index in milk
From Table 10, with LCGroup comparison, LEPThe group improves the T-AOC antioxidant capacity of 21-day-old milk (P<0.05），LPGroup has no significant effect: (P>0.05), but LPGroup had tendency to increase T-AOC in 28-day-old milk (P=0.066），LEPGroup has no significant effect: (P>0.05）。 LPGroup LEPGroup sum had no significant effect on T-AOC in 14 day old milk (P>0.05). And LCGroup comparison, LPThe group increased the MDA content in 21-day-old milk ()P<0.05），LEPGroup has no significant effect: (P>0.05), has no significant effect on the MDA content in 14 and 28 day old milk (P>0.05）。LPGroup sum LEPGroup 14, 21 and 28 day old milksThe enzyme activities of GSH-PX, SOD and CAT have no significant influence (P>0.05）。
TABLE 10 Effect of perinatal Nicotinamide addition on antioxidant levels in milk
1LC= control, LP= group around postpartum period, LEP= perinatal full period group.
The difference of the lower case letters on the same row of shoulder marks is significant (P<0.05）。
2.2.5 serum antioxidant index
From Table 11, with LCGroup comparison, LEPGroup significantly reduced serum TAOC in 14 day old lambs (P<0.05），LPGroup has no significant effect: (P>0.05), but LPGroup significantly reduced serum TAOC in 28-day-old lambs (P<0.05），LEPGroup has no significant effect: (P>0.05）。LEPGroup significantly reduced CAT enzyme activity in serum from 28-day-old lambs compared to control group (P<0.05), has no significant effect on CAT enzyme activity in 14-day-old serum (P>0.05），LPNo significant effect on CAT enzyme activity in serum of 14-day and 28-day-old lambs (groupP>0.05）。LPGroup sum LEPHas no significant influence on the GSH enzyme activity, the SOD enzyme activity and the MDA content of the serum of 14-day and 28-day-old lambs ()P>0.05）。
TABLE 11 influence of Nicotinamide addition on lamb serum antioxidant index in perinatal period
1LC= control, LP= group around postpartum period, LEP= perinatal full period group.
The difference of the lower case letters on the same row of shoulder marks is significant (P<0.05）。
2.2.6 liver antioxidant index
From Table 12, with LCGroup comparison, LEPGroup significantly reduced CAT enzyme activity in liver tissue of 28-day-old lambs: (P<0.05），LPGroup has no significant effect: (P>0.05）。LPGroup and LEP group had no significant effect on TAOC and SOD enzyme activity in liver tissue of lamb ((P>0.05）。
TABLE 12 influence of Nicotinamide addition on lamb liver antioxidant index in perinatal period
1LC= control, LP= group around postpartum period, LEP= perinatal full period group.
The difference of the lower case letters on the same row of shoulder marks is significant (P<0.05）。
2.2.7 relative expression level of liver antioxidase mRNA
As can be seen in FIG. 3, and LCGroup comparison, LPGroup sum LEPThe group significantly reduced the mRNA expression levels of SOD2 and GPX1 in liver tissues of 28-day-old lambs (a)P<0.05）。LEPSignificantly reduced CAT mRNA expression in liver tissue of lambs compared to control group (P<0.05），LPGroup has no significant effect: (P>0.05）。LPGroup sum LEPNo significant influence is caused on the mRNA expression quantity of GPX2 and GPX3 in liver tissues of 28-day-old lambs: (P>0.05）。
2.3 Effect of Nicotinamide addition to Dairy goats in perinatal period on lamb immune function
2.3.1 immunological indicators in colostrum
From Table 13, with LCGroup comparison, LPGroup sum LEPGroup has tendency to decrease IL-6 content in colostrum: (P= 0.081), but has no effect on the concentration of immunoglobulin (P>0.05）。
TABLE 13 influence of immune-related indices in Nicotinamide colostrum addition to ewes in perinatal period
2.3.2 immunological indices in Normal milk
From Table 14, with LCGroup comparison, with LEPGroup reduced IL-1. beta. content in 28 days milk ((S))P<0.05），LPGroup has a decreasing tendency (P= 0.062), had no significant effect on IgG concentration in 14, 21, and 28 days milk: (P>0.05）。
TABLE 14 influence of Nicotinamide addition to ewes in perinatal period on immune-related indices in colostrum
2.3.4 serum protein content
From Table 15, with LCGroup comparison, LPGroup sum LEPThe group had no significant effect on total protein, albumin, globulin and white-to-globulin ratios in serum of 14-day-old and 28-day-old lambs (P>0.05）。
TABLE 15 Effect of Nicotinamide addition on lamb serum protein content in perinatal period
2.3.5 immune-related indicators in serum
From Table 16, with LCGroup comparison, LPGroup sum LEPThe group improves the content of IgG in the serum of 28-day-old lambs (P<0.05), has no significant effect on the IgG content in the serum of 14-day-old lambs (P>0.05）。LPGroup and LEP group there was no significant effect on IL-6 and TNF- α concentrations in serum of 14 and 28 day old lambs.
TABLE 16 Effect of peripartum Nicotinamide addition on serum immune markers
2.3.6 spleen cytokine mRNA expression level
As can be seen in FIG. 4, and LCGroup comparison, LPGroup significantly improved mRNA expression level of IL-1 beta in spleen tissue of 28-day-old lamb ((P<0.05），LEPGroup has a tendency to increase (P=0.062）。LPGroup sum LEPmRNA expression of IL-6, IL-8 and TNF- α in spleen tissue of group lambs had no significant effect (II)P>0.05）。
As can be seen in FIG. 5, and LCGroup comparison, LEPGroup significantly reduced IFN-gamma mRNA expression level in spleen tissue of 28-day-old lamb ((P<0.05），LPGroup has no significant effect: (P>0.05）。LPGroup sum LEPNo significant effect on the mRNA expression level of IL-2 and IL-4 in spleen tissue of lamb group: (P>0.05）。
2.3.7 mesenteric lymph node cytokine expression level
As can be seen in FIG. 6, and LCGroup comparison, LPGroup sum LEPThe group obviously improves the mRNA expression quantity of IL-1 beta in lymph node tissues of 28-day-old lambs ((P<0.05）。LPGroup sum LEPThe group has no significant influence on the mRNA expression quantity of proinflammatory cytokines in IL-6, IL-8 and TNF-alpha in lamb lymphoid tissues (P>0.05）。
As can be seen in FIG. 7, and LCGroup comparison, LEPGroup significantly reduced the mRNA expression levels of IL-2 and IFN-gamma in lymphoid tissues of 28-day-old lambs ((P<0.05），LPNo significant effect on the mRNA expression level of IL-2 and IFN-gamma (P>0.05）。LPGroup sum LEPNo significant effect on the expression level of IL-4 mRNA in lymphoid tissues of 28-day-old lambs: (P>0.05）。
2.3.8 relative expression level of DNMTs mRNA
As shown in FIG. 8, with LCGroup comparison, LPGroup sum LEPThe group improves the mRNA expression quantity of the lamb spleen tissue DNMT1 (P<0.05), but had no effect on the mRNA expression level of DNMT3b (P>0.05）。
3 conclusion
(1) The nicotinamide added to the ewes in the late perinatal period and the whole period has no influence on the birth weight of the lambs and the growth and development of the ewes at the age of 28 days, and also has no influence on the gestational period of the ewes.
(2) Nicotinamide is added to ewes in the full perinatal period to reduce the activity of T-AOC and CAT enzymes in plasma of 14-day-old lambs, and nicotinamide is added in the late perinatal period to reduce the T-AOC in plasma of 28-day-old lambs. Results in liver tissues show that the enzyme activity of CAT in the tissues is reduced by adding nicotinamide in the perinatal period, the mRNA expression levels of SOD2 and GPX1 in the tissues are reduced by adding nicotinamide in the perinatal period and the perinatal period, and the mRNA expression level of CAT is reduced by adding nicotinamide in the perinatal period, which indicates that the oxidative stress state of lambs is reduced by adding nicotinamide in the perinatal period and the perinatal period of ewes. The perinatal period of the dairy animals is in a special stage, the metabolism intensity is high, the oxygen consumption is high, and a large amount of active oxygen free radicals are easy to generate. These reactive oxygen radicals are transmitted to the fetus before birth of the lamb via the placenta and to the breast milk after birth. The antioxidant system of the body is mainly used for eliminating active free radicals, and the activity of antioxidant enzyme is related to the concentration of active oxygen free radicals. Nicotinamide improves the antioxidant capacity of the mother body and milk, and reduces the transmission of active free radicals to offspring through placenta and milk, thereby reducing the oxidative stress of lambs.
Thus, it is known that the addition of niacinamide to the full-term perinatal ewes reduces the transfer of reactive oxygen radicals to the fetus via the placenta. The nicotinamide is added to the ewes in the late perinatal period and the whole period, so that the oxidation resistance of milk is improved, the active oxygen free radicals are reduced to be transmitted to offspring through the milk, and the oxidative stress of lambs is reduced.
(3) The addition of nicotinamide to the ewes in the late perinatal period and the full period reduces the gene expression of lamb spleen DNMT1, but has no influence on the expression of DNMT3b, and shows that the lamb spleen DNA methylation level tends to be reduced. Nicotinamide N-methyltransferase catalyzes nicotinamide methylation by taking S-adenosylmethionine as a methyl donor, and is an important methyltransferase in animal bodies. The addition of nicotinamide may competitively deplete the body of methyl groups, while protein poly (ADP-ribose) polymerases (PARPs) and sirtuin deacetylases, which rely on Nicotinamide Adenine Dinucleotide (NAD), play an important role between cellular metabolism and gene expression influenced by various pathways (modification of DNA and histones). Low levels of NAD promote BDNF activity dependent promoter DNA methylation.
Therefore, the addition of nicotinamide to the ewes in the late perinatal period and the full period improves the immune function of the lambs, and the improvement of the immune function may be related to the change of the methylation level of DNA.
The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments, and it should be understood by those skilled in the art that various modifications or changes can be made by those skilled in the art without inventive efforts based on the technical solutions of the present invention.
Claims (1)
1. A method for enhancing the antioxidant and immune functions of lambs by adding nicotinamide to dairy goats in the perinatal period is characterized in that the perinatal period comprises the late gestation period and the early lactation period of the dairy goats; the perinatal period is from 21 days before delivery to 28 days after delivery of the milk goat;
the nicotinamide addition amount is 5 g of nicotinamide fed to each perinatal milk goat every day;
the adding method is carried out by adopting a direct drenching mode.
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