Pregnancy toxaemia (ketosis) is a metabolic disease of ewes which occurs during the late gestation as a result of the inability of the pregnant ewe to maintain an adequate energy balance for the fast growing maternal fetal unit. As a result of energy defi-ciency mobilization of lipid reserves results in a doubling of the plasma free fatty acid (FFA) giving rise to fatty liver and increased ketone bodies β-hydroxybutyrate (BHB) in blood and urine. It is associated with a higher rate of mortality and causes severe economic losses.
The objective of this study was directed at investigating the relationship between metabolic parameters and cytokine TNFα, to check the interaction between the TNFα and fat metabolism in late pregnant ewes of different breeds, and whether TNFα play a role in the pathogenesis of pregnancy toxaemia, which may serve as marker to early diagnosis of the disease.
In this study, 29 pregnant and clinically healthy ewes (16 Merino, 13 Blackhead) were selected out of a flock of sheep. Blood samples were collected at 5, 3, and 1 week be-fore parturition (b.p.) and also 4 weeks after parturition (a.p.). The average numbers of lambs were 2.18 and 1.58 /ewe for Merino and Blackhead breeds respectively. The blood samples were analyzed for the following:
Concentration of metabolic parameters: glucose, insulin, free fatty acids (FFA), β-hydroxybutyrate (BHB), albumin, total protein (TP), iron (Fe), glutamat-dehydro-genase (GLDH), creatin kinase (CK), gamma-glutamyl-transferase (GGT), choles-terol, haptoglobin.
Haematological parameters: Haematocrite (HK), haemoglobin concentration (HB), erythrocyte count (EC), leukocyte count (LC), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC).
Cytokine TNFα by using ovine TNFα ELISA assay.
The results of glucose concentration of pregnant ewes showed significant increase (3.8 mmol/l) in five weeks b.p. and declined with advancing gestation (2.6 mmol/l) one week b.p. Insulin concentration remained constant with an average of 0.11 nmol/l b.p., and then significantly increased to 0.22 nmol/l four weeks a.p.
Maximal FFA concentrations were found at five weeks b.p. (976 µmol/l). The levels of FFA showed high levels b.p. compared with reference range (R.R. < 600 µmol/l), and the FFA levels significantly decreased postpartum (four weeks b.p.). while there was significant increasing (p<0.05) in the level of FFA in Merino sheep than in Black-head sheep b.p. On the other hand there was no significant difference a.p.
The mean values of BHB in all periods of sampling, period 1(5 w.b.p.), period 2 (3 w.b.p.), period 3 (1 w.b.p.), and period 4 (4 w.a.p.) were 0.37 mmol/l, 0.23 mmol/l, 0.17 mmol/l and 0.3 mmol/l respectively. The mean of BHB indicated normal levels of BHB before and after parturition compared to subclinical ketosis (BHB > 1 mmol/l) and clinical ketoses (BHB > 1.6 mmol/l), and there was a significant difference (p<0.05) in the values of BHB between Blackhead and Merino breeds before parturi-tion while there was no significant difference after parturition.
The concentration of TNFα showed elevated levels in all period of sampling before parturition. The TNFα values were 30.4 (17.2, 785.0) ng/ml (median, first, and third quartiles), 35.6 (13.6, 54.3), and 26.6 (13.0, 39.9) ng/ml in period 1(5 w.b.p.), period 2 (3 w.b.p.), and period 3 (1 w.b.p.) respectively. These values decreased to 19.1 (9.9, 33.8) ng/ml at 4 weeks after parturition.
Statistical analysis showed that there was a positive correlation between free fatty ac-ids and TNFα. This correlation means that adipose tissue produces TNFα causing insu-lin resistance, which stimulates the lipolysis and leads to an increase of circulatory free fatty acids levels.
It is concluded that fat mobilization occurs in the prepartum clinically healthy ewes with a significant increase in the levels of FFA, and also there is an increase in the proinflammatory cytokine TNFα at late gestation which predisposes ewes to pregnancy toxaemia and can aid in the diagnosis of the disease.:Table of contents
1 Introduction…………….…………………………………………………..1
2 Review of literature……....…………….……………………………….….3
2.1 Metabolic condition of ewes in the late pregnancy………….………………3
2.2 Metabolic disorders subacute and acute pregnancy Toxaemia…….………..4
2.3 The Importance and dynamic of the different biochemical parameters
glucose, insulin, free fatty acids, and β-hydroxybutyrate of ewes in the
late pregnancy…………………….………..….……………………………10
2.3.1 Glucose……………………………………………………………………..10
2.3.2 Insulin………………………………………………………………………12
2.3.3 β-hydroxybutyrate (BHB)…….……………………………………………16
2.3.4 Free fatty acids……………………………………………………………...20
2.4 TNFα and its role in fat metabolism and pregnancy toxaemia………….….22
3 Animals, material and methods……………………………………..……38
3.1 Animals……………………………………………………………………..38
3.2 Clinical examination………………………………………………………..38
3.3 Collection of blood samples……………………………………………..…38
3.4 Analysis of haematological parameters in blood samples……………….....38
3.5 Determination of biochemical parameters………………………………….39
3.6 Determination of haptoglobin by using haptoglobin assay……………...…39
3.6.1 Haptoglobin assay principle………………………………………………..39
3.6.2 Components………………………………………………………………...40
3.6.3 Additional materials required………………………………………………40
3.6.4 Sample and reagent preparation………………………………….….……..40
3.6.4.1 Samples……………………………………………………………………..40
3.6.4.2 Haemoglobin.................................................................................................40
3.6.4.3 Chromogen/Substrate………………………………………………………40
3.6.5 Manual methods (microplate or spectrophotometric)……………………...41
3.6.5.1 Calibrator…………………………………………………………………...41
3.6.5.2 Test temperature…………………………………………………………....41
3.6.5.3 Procedure………………..………………………………………………….41
3.7 Analysis of TNFα by using ovine TNFα ELISA assay…………………….42
3.8 Statistical analysis…………………………………………………….…….43
4 Results.……………………………………………………………………..44
4.1 Clinical examination (observation)…………………………………….…...44
4.2 Biochemical parameters………………………………………………….…44
4.2.1 Glucose………………………………………………………………….…..44
4.2.1.1 Glucose concentrations in all sheep…………………………..…………….44
4.2.1.2 Glucose concentrations in Blackhead sheep…………………………..…....44
4.2.1.3 Glucose concentrations in Merino sheep………………………...…………45
4.2.2 Insulin……………………………………………………………………….45
4.2.2.1 Insulin concentrations in all sheep………………………………..………...45
4.2.2.2 Insulin concentrations in Blackhead sheep………………………..………..46
4.2.2.3 Insulin concentrations in Merino Sheep…………………………………….46
4.2.3 Free fatty acids……………………………………………………………...47
4.2.3.1 Free fatty acid concentrations in all sheep………………………………….47
4.2.3.2 Free fatty acid concentrations in Blackhead sheep………………………....47
4.2.3.3 Free fatty acid concentrations in Merino sheep………………………….....47
4.2.4 β-hydroxybutyrate (BHB)…….…………………………………………….48
4.2.4.1 β-hydroxybutyrate concentrations in all sheep……..……………………....48
4.2.4.2 β-hydroxybutyrate concentrations in Blackhead sheep……… …………....48
4.2.4.3 β-hydroxybutyrate concentrations in Merino sheep…………………...…...49
4.2.5 Tumor necrosis factor alpha (TNFα)………………….……………….…...50
4.2.5.1 TNFα concentrations in all sheep…………..................................................50
4.2.5.2 TNFα concentrations in Blackhead sheep……………………………….....50
4.2.5.3 TNFα concentrations in Merino sheep………………………………….......51
4.2.6 Haptoglobin………………………………………………………………....51
4.2.6.1 Haptoglobin concentrations in all sheep……………………………..…......51
4.2.6.2 Haptoglobin concentrations in Blackhead and Merino sheep…….………..51
4.2.7 Albumin…………………………………………………………………….52
4.2.7.1 Albumin concentrations in all sheep…………………………..…………...52
4.2.7.2 Albumin concentrations in Blackhead and Merino sheep……………….....53
4.2.8 Creatinkinase (CK)…………………………………………………………53
4.2.8.1 Creatinkinase activity in all sheep………………………………………….53
4.2.8.2 Creatinkinase activity in Blackhead and Merino sheep…………………….53
4.2.9 Gamma-Glutamyl Transferase (GGT)………………………….…………..54
4.2.9.1 GGT activity in all sheep…………………………………………………...54
4.2.9.2 GGT activity in Blackhead and Merino sheep……………………………..55
4.2.10 Glutamat-Dehydrogenase (GLDH)………………………………………...56
4.2.10.1 GLDH activity in all sheep…………………………………………………56
4.2.10.2 GLDH activity in Blackhead and Merino sheep………….…………….......56
4.2.11 Total protein………………………………………………………………..57
4.2.11.1 Total protein concentrations in all sheep………………………………...…57
4.2.11.2 Total protein concentrations in Blackhead and Merino sheep……………..57
4.2.12 Cholesterol……………………………………………………………….…58
4.2.12.1 Cholesterol concentrations in all sheep………………………………...…...58
4.2.12.2 Cholesterol concentrations in Blackhead and Merino sheep…………....….58
4.2.13 Iron………………………………………………………………………….59
4.2.13.1 Iron concentrations in all sheep…………………………...………………..59
4.2.13.2 Iron concentrations in Blackhead and Merino sheep………………….........60
4.3 Haematological parameters………………………………………………...60
4.3.1 Haematological parameters in all sheep……………………………………60
4.3.2 Haematological parameters in Blackhead sheep…………………………...61
4.3.3 Haematological parameters in Merino sheep………………………………61
4.3.4 Statistical analysis of haematological parameters………………………….62
4.3.4.1 Haemoglobin concentration (Hb)…………………………………………..62
4.3.4.2 Haematocrite (HK)…………………………………………………………62
4.3.4.3 Mean corpuscular volume (MCV) …………………………………………62
4.3.4.4 Mean corpuscular haemoglobin (MCH)……………………………………62
4.3.4.5 Mean corpuscular hemoglobin concentration (MCHC)……………………63
4.3.4.6 Thrombocytes volume (THB)……………………..…………………….…63
4.3.4.7 Leukocytes (LC)………..…………………………………………………..63
4.3.4.8 Erythrocytes.……………………………………………………………….63
5 Discussion…………………………………………………………………..65
6 Summary…………………………………………………………………..71
7 Zusammenfassung………………………………………………………...73
8 References…………………………………………………………………75
Acknowledgements.......................................................................................................84
Identifer | oai:union.ndltd.org:DRESDEN/oai:qucosa:de:qucosa:11212 |
Date | 06 July 2011 |
Creators | El-Ebissy, Eman |
Contributors | Fürll, Manfred, Ganter, Martin, Universität Leipzig |
Source Sets | Hochschulschriftenserver (HSSS) der SLUB Dresden |
Language | English |
Detected Language | English |
Type | doc-type:doctoralThesis, info:eu-repo/semantics/doctoralThesis, doc-type:Text |
Rights | info:eu-repo/semantics/openAccess |
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