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Comparative biochemical indices of fetal fluids in normal foaling and stressful delivery in Indian thoroughbred mares
Refereed
COMPARATIVE BIOCHEMICAL INDICES OF FETAL FLUIDS IN NORMAL FOALING AND STRESSFUL DELIVERY IN INDIAN THOROUGHBRED MARES H.P.S. Kochhar, MVScl; P.S. Simran, MVSc2; A.S. Nanda, FRCVS, PhD1; Ripudaman Kaur, MVSc
SUMMARY
This study was conducted on 22 mares for establishing the vital biochemical parameters in the amniotic fluid and allantoic fluid of mares having normal parturition and difficult/stressful foaling (dystocia). Evaluation of the biochemical constituents in fetal fluids revealed that other than bilirubin, alkaline and acid phosphatase and lactate dehydrogenase, none of the biochemical parameter varied significantly amongst the two groups of normal foaling and dystocia suffering mares. The significant variation in the values of transaminases, estrases, creatine, urea nitrogen, glucose, sodium, potassium, calcium and phosphorus amongst the amniotic and allantoic fluids of each group (normal and dystocia) was observed. Variation in the biochemical indices could be attributed to fetal struggle and stress to the mare during dystocia. The allantoic and amniotic fluids vary widely in terms of biochemical constituents in each group.
INTRODUCTION
The volume of fetal fluids in mares, which is about 1020 liters towards term is regulated mainly by swallowing and excretion by the fetus. 1 The viscous amniotic fluid is derived from skin, mucous, amniotic epithelium fetal saAuthors' address:
1Department of Gynaecology & Obstetrics;
2Department of Surgery & Radiology, College of Veterinary Sciences, Punjab Agricultural University, Ludhiana-141 004 India. Acknowledgments: Authors wish to sincerely thank M/S Dasmesh Stud Farm, Sarai Naga, Faridkot, Punjab, India for providing the facilities.
206
liva and naso-pharyngeal secretions, while allantoic fluid primarily consists of excretion of fetal kidneys. 2 These fluids contain varying amounts of enzymes, phospholipids, sugars and ions, which are produced by the fetus. 3 The study of various biochemical constituents of fetal fluids could therefore serve as an index of fetal health even ante partum. Studies have been conducted on composition of fetal fluids in normal foaling 4 or in mid gestation, t The present study was planned with a view to establish the concentration of various enzymes and other vital constituents in fetal fluids of mares foaling normally or with assistance, i.e., dystocia. Efforts were made to correlate concentrations in both the groups, and establish variation if any in gestation length, fetal weight and placental weight in normal and difficult foaling mares.
MATERIALS AND METHODS
This study was conducted in the foaling season at an organized stud farm. In all 22 mares were taken into study, 11 having normal foaling and 11 requiring assistance up to moderate fetal mutations during foaling. The allantoic and amniotic fluids were collected by puncturing the respective sacs when presented in the birth canal, with the help of a sterile needle and syringe. Gestation length, parity, fetal sex and fetal weight, and placental weight were also recorded. The samples were transported on ice to the laboratory and were frozen at -20~ pending analysis within 3-5 days. The concentration of various biochemical constituents viz. glucose, 5 urea nitrogen, 6cholesterol, 7 aspartate aminotransferase 8 (AST), alanine aminotransferase 8 (ALT), alkaline JOURNAL OF EQUINE VETERINARY SCIENCE
phosphatase a (AKP), acid phosphatase a (ACP), acetylcholinesterase 9 (ACHE), butryl cholinesterase 10 (Butryl ChE), carboxylestrase 01 (Crb E), arylesterase 12 (ARE), lactate dehydrogenase 8 (LDH) and phosphorus la were determined by standard methods. Total proteins, albumin, bilirubin, creatinine were measured by standard kits. a Sodium (Na), potassium (K) and calcium (Ca) were estimated using Atomic Absorption Spectrophotometer. b Paired 't' test was employed for statistical analysis of the data. TM
Table 1. P r e g n a n c y d a t a of n o r m a l f o a l i n g a n d d y s t o c i a suffering m a r e s . Dystocia n = 11
Normal foaling n = 11
Gestation period (days)
332+7.79
323.87+7.9
Age of mares
9.4+3.61
11.13+4.83
3+1.18
4.37+2.86
Foal wt. (Kg)
53.5+7.09
50+7.07
Placenta wt. (Kg)
5.3+ 1.18
4.75+0.43
5 :6
6 :5
No. of mares
(yrs) Parity
RESULTS
All the foals delivered were live and various observations of normal and dystocia affected mares is tabulated in Table 1. The foal and placental weights, gestation length and age of mares were not statistically different between the two groups. The allantoic fluid was of amber color, clear and watery in consistency while the amniotic fluid was whitish, viscid and transopaque. The fetal fluids were free from blood, meconium or intermixing. The concentration of AKP, ACD, AST, ALT, ACHE, Butryl ChE, Carb E, Aryl E, urea nitrogen and glucose were significantly higher in amniotic than in allantoic fluid (p < 0.05) of mares both with normal as well as with assisted delivery. On the contrary concentration of LDH, globulin, bilirubin and creatinine were significantly higher (p < 0.05) in allantoic fluid than in amniotic fluid of both normal and dystocia suffering mares. The concentration of total proteins, albumin, immunoglobulin and cholesterol were similar in both types of fetal fluids. The concentration of AST, ALT butyl ChE, Carb E, Aryl E, total proteins, globulins, albumin, creatinine, glucose and cholesterol in allantoic fluid of dystocia affected mares were not significantly different than those in the normal mares. However, a significantly higher than normal (p < 0.05 & p < 0.001) concentration of AKP, LDH, bilirubin and BUN and lower concentration of ACP were observed in allantoic fluid of dystocia affected mares. The amniotic fluid concentration of bilirubin and AKP were significantly higher (p < 0.05) in mares with dystocia as compared to normal foaling ones.
DISCUSSION
The fetoplacental unit is a dynamic system with a constant exchange between fetal fluid compartments and maternal circulation. 2 A variation in the concentration of various biochemical components of allantoic and amniotic fluids could have a significant relation to the fetal health and may indicate certain fetal diseases, cystic fibrosis and aKits: Spans Diagnostics Pvt. Ltd. 173-B, New Industrial Estate, Udhana (Surat), India. bAtomic Absorption Spectrophotometer (Model AA6) Varian Techtran, Melbourne, Australia
Volume 17, Number 4, 1997
Sex ratio (Male : Female)
other conditions described in humans. 15 The alkaline and acid phosphatases are correlated with fetal maturity, fetal bone formation and kidney functioning, t In early pregnancy the AKP activity is mainly of fetal intestine origin. Meconium release or fetal defecation in distress contributes to its increased activity. Presumably AKP activity represents entry of desquamated intestinal mucosal cells 16 into fetal fluids. The concentration of AKP was significantly increased both in allantoic and amniotic fluid of mares suffering from difficulty at parturition as compared to their normal counterparts. The values in normally foaling mares corresponded with earlier studies. 1No reference for comparison of studies in dystocia affected mares are available for confirmation. The ALT and AST values are indicative of fetal liver functioning and muscular injuries if any. Their significantly different values for the amniotic and allantoic fluid are suggestive of the nature of fluid in each compartment, i.e., intestinal and tracheal secretions in amniotic fluid and fetal urine in allantoic fluid. 17 The concentration of esterases viz. ACHE, Butryl ChE, Carb E, Aryl E in amniotic and allantoic fluids of dystocia affected mares was significantly (p < 0.05) lower than normal. Although AChE is generally associated with neural tube defects or defects leading to leakage of CSF during fetal formation; however, it has rarely been observed in normal fetuses 1s,t9 too. Information on the source of these enzymes in fetal fluids is obscure. Perhaps stress due to enhanced fetal moments or struggle during dystocia might be a contributing factor to the concentration of esterases amongst the two groups. Lactate dehydrogenase is contributed by all types of cells including exfoliative cells. 1 Its rise has been reported with advance in fetal age and fetal death, a The rise in LDH, in the fetal fluids of dystocia suffering mares could be attributed to enhanced cellular exfoliation, escape of fetal intestine contents or due to extra fetal muscular movements during struggle to 207
Table 2 . B i o c h e m i c a l i n d i c e s in fetal f l u i d s of n o r m a l f o a l i n g a n d d y s t o c i a s u f f e r i n g m a r e s . Parameter
Units
AKpwwxxyYzz
IU/L
25.63+2.38
ACP wwxxyY
IU/L
1.91 +0.8
AST xwx
IU/L
ALT wx
lU/L
LDH wxxyY
I U/L
122.3_+14.5
AChE wxy
nmol thiol formed min-1 ml-1
14.94_+14.14
4.22+3.12
26.40+2.28
21.50_+22.72
Butryl ChE wwxx
nmol thiol formed min-1 ml-1
5.41 _+4.75
2.54_+2.06
13.49_+4.31
13.40_+7.26
Carboxyl Est wxx
nmol indophenol formed min-1 ml-1
11.25_+3.93
12.22_+3.77
25.00_+18.78
22.78_+9.36
nmol phenol formed min-1
0.09_+0.02
0.13_+0.07
0.69-+0.92
0.02_+0.004
UN wwy
mg/dL
27.75_+9.48
40.17_+8.16
41.7_+7.2
48.0_+9.0
Total protein x
gm/dL
1.21_+0.34
1.40_+0.31
0.91_+0.2
0.97_+0.37
Globulin wxx
grn/dL
1.02_+0.2
1.18_+0.14
0.78_+0.12
0.82_+0.2
Albumin
gm/dL
0.08_+0.3
0.13_+0.5
0.04_+0.06
0.05_+0.3
Total BilirubinWWxxyYzZ
mg/dL
1.2_+0.5
1.85_+0.23
0.21_+0.12
0.85_+0.32
Creatinine wWxX
mg/dL
68.7_+14.9
72.3_+8.8
19.38_+4.67
20.14_+4.77
Immunoglobulins
g/dL
0.87_+0.32
0.93_+0.56
0.63_+0.28
0.72_+0.34
Glucose wwxx
mg/dL
5.8_+1.19
6.2_+3.4
20.1_+5.4
21.6_+8.2
Cholesterol
mg/dL
8.8_+1.2
7.9_+3.4
7.2_+2.6
6.8_+3.1
Sodium wwxx
meq/L
71 _+23.8
88_+20.1
146_+4.8
149_+6.2
Potassium wwxx
meq/L
26.2_+10.8
28.4_+8.0
9.2_+1.8
10.4_+2.6
Calcium wwxx
meq/L
31.2_+9.8
34.3_+6.6
11.6_+2.8
12.2_+8.6
Phosphorus wwxx
meq/L
22.3_+8,7
26.6_+11.3
4.0_+1.8
4.4_+2.2
Aryl Est wxxzz
1) Normal Foaling:
Allantoic fluid vs Amniotic fluid w - differs significantly at p < 0,05 W - differs significantly at p < 0.001
Allantoic fluid Normal Dystocia 34.36+4.61
96.95+10.15
103.23_+13.11
1.3+0.4
6.59+1.51
8.44_+2.85
6.94_+3.38
7.48_+4.45
14.01 _+3.63
17.02_+10.91
9.70_+7.52
9.09_+7.99
24.78+13.94
16.92_+10.86
98.17_+ 18.2
112.8+21.4
178.0_+52.4
ml-1
3) Allantoic fluid: 4) Amniotic fluid:
2) Dystocia suffering mares: Allantoic vs Amniotic x - differs significantly at p < 0.05 X - differs significantly at p < 0.001
208
Amniotic fluid Normal Dystocia
Normal vs Dystocia suffering y - differs significantly at p < 0.05 Y - differs significantly at p < 0.001 Normal vs Dystocia z - differs significantly at p < 0.05 Z - differs significantly at p < 0.001
JOURNAL OF EQUINE VETERINARY SCIENCE
take birth. In dystocia affected animals the amniotic protein values were relatively lower than normal which is similar to that reported in bovines, a Concentration of bilirubin, a product of bile is indicative of liver health of animals. Enhanced biliary secretions may be the cause of increased bilirubin in both types of fetal fluids of dystocia suffering mares. In humans it has been reported that amniotic fluid bilirubin concentration becomes higher than in the maternal serum after 34 weeks of pregnancy probably due to decreased excretory ability of the maternal liver. 2~ Fetal urine contributes significantly to the allantoic fluid. 1 This may explain the rise of urea nitrogen and creatinine in stressfully foaling mares because of increased fetal micturition during violent efforts at the time of birth. Also in pregnancy creatinine moves from mother to fetal serum and then to fetal urine and to anmiotic fluid. 21 At times, however, creatinine may originate from fetal muscles and increased protein metabolism, a~ The glucose from maternal circulation is the main energy source for the fetus during normal pregnancy in ovines, aa although some of it is reduced to sorbitol in the placenta, a3 Our findings of lower glucose in the allantoic than amniotic fluid at foaling are in conjunction with those in bovines. 24 Increased glucose concentration in amniotic fluid has also been taken as an index of infectious state in pre-term labor in women, as Absence of any change from normal values in dystocia mares suggest it to be free from infections. It has been established that fetal renal excretion is the probable origin of free fatty acid in amniotic fluids of women. 26 The cholesterol represents one-third of total lipid profile in amniotic fluid but then it remains unchanged despite increase in quantity of amniotic fluid nearing term. 27 Immunologic activity of amniotic fluid remains poorly understood. 17 Allantoic fluid had much lower concentration of sodium than in amniotic fluid suggesting that the latter closely resembles the composition of plasma transudate and the flow of Na + from serum of mother to amniotic fluid is easy. 28'1 The high concentration of potassium in allantoic fluid is suggestive of increased fetal urine and metabolic activity of the fetus at tenn. Potassium ions differ in biological activity from sodium, as they cannot undergo passive transfer across cell membranes.29 High concentration of calcium in allantoic fluid is again attributed to fetal kidney status. The different biochemical constituents established in the above study are an effort to report the variation due to different foaling processes. Stress of dystocia, fetal movements in distress and forceful expulsive efforts by the mother may have contributed to variations in the values of some vital parameters. The endocrine studies may be correlated with such indices in an effort to bring forth the effect of stress of dystocia on biochemical constitution of fetal fluids in mares. Volume 17, Number 4, 1997
REFERENCES 1. Williams MA, Wallace Ss, Tyler JW, McCall CA, Gutierrez A, Spano JS; Biochemical characteristics of amniotic and allantoic fluid in late gestational mares. Theriogenology 1993;40:1251257. 2. Baetz AL, Hubert WT, Graham CK: Changes of biochemical constituents in Bovine fetal fluids with gestoterial age. Am J Vet. Res 1976;37:1047-1052. 3. Mohamed AR, Noakes DE: Enzyme activities in amniotic fluid and maternal blood in cattle before and after induced foetal death and abortion. Br Vet J 1985;141 (1):49-59. 4. Schott HC, Mannsman RA: Biochemical profiles of normal equine amniotic fluid at parturition. Equine Vet J 1988;5:52. 5. Frankel S, Reitman S, Sonnenwirth AC: In: Gradwohl's Clinical Laboratory Methods and Diagnosis. 7th edn. Vol. 1 St. Louis, USA: CV Mosby Company 1970. 6. Evans RT: Manual and automated methods of measuring urea based on a modification of its reactions with diacetyl monoxine and thiosemicarbazide. J Clin Pathol 1968;21:527. 7. Zak B: Simple and rapid micro technique for serum total cholesterol. Am J Cl in Pathol 1957;28:583. 8. Wooton IDP: Microanalysis in Medical Biochemistry. 4th edition. J&A Churchill Ltd. London UK, 1964. 9. Morari K, Usbiyama S, Satoh T, KugaT: Enzyme induction by repeated adminstration of tetrachlorvinophos in rats. Toxicol Appl Pharmacol 1976;37:377. 10. Vors G, Sachsso K: Red cell and plasma cholinestrase activities in micro samples of human and animal blood determined simultaneously by a modified acetylcholine/DTNB procedure. Toxicol Appl Pharmacol 1970;16:764. 11. Mendoza CE, Shields JB, Phillips WE J: Distribution of carboxylesterase activities in different tissues of albino rats. Comp Biochem Physiol 1971 ;40:841. 12. Mc Ilvain JE, Timoszyk J, Nakatsugawa T: Rat liver paraxonase (paraxona arylesterase). Pesticide Bioch and Physiol 1984;21:162. 13. Verlag H: Practical Clinical Biochemistry. 5th edn. William Hienemann Medical Books Ltd. London, 1980. 14. Snedecor GW, Cochran WG: Statistical methods 8th edition. Calcutta, India: Oxford and IBH Publishing Co. 1967. 15. Szabo M, Veress L, Teichmann F, Munich A, Huszka M, Papp Z; Amniotic fluid microviilar enzyme activity in fetal malformations. Clin Genet 1990;38:340-345. 16. Hahnemann N, Sorenson SA: Studies on alkaline phosphatase in amniotic fluid. Acta Obstet Gynaecol Scand 1974; 53:15-22. 17. Gleicher N: Immunological influence of amniotic fluid. Lancet 1980;1:541. 18. Barlow RD, Cuckle HS, Wald NJ: False positive gel acetylcholinestrase results in blood stained amniotic fluids. Br J Obstet Gynaecol 1982;89:821-829. 19. Goldfine C, Hadow J, Hudson G: Densitometry as an aid in amniotic fluid gel acetylcholinestrase analysis. Am J Obstet Gynaecol 1983; 145:317. 20. Benzie RJ, Doran TA, Harkins JL, Owen VMS, Parker CJ: Composition of amniotic fluid and maternal serum in pregnancy. Am J Obstet Gynaecol 1974;119:798. 21. Hodari AA, Mariona FG, Houliham RT: Creatinine transport in the maternal fetal complex. Obset Gynaecol 1973;41 : 47. 22. Alexander DP, Britton HG, Nixon DA: The metabolism of fructose and glucose by the sheep foetus: Studies on isolated perfused preparation with radioactivity labeled sugars. J Exp Med 1970;55:346-362. 23. Bdtton HG, Hugget ASG, Nixon DA: Carbohydrate metabolism in sheep placenta. Biochem BiophysActa 1967;136: 426-440. 24. Reeves JT, Daoud MD, Gentry M, Eastin C: Changes in
209
urinary flow in bovine fetuses during late gestation: Comparison of amniotic and fetal body fluids. Am J Vet Res 1972;33(11 ):21592167. 25. Romero R, Jimenez C, Lohda AK, Neres J, Hanaoka S, Avila C, Callahan R, Mazoi M, Hobbins JC, Diamond MP: Amniotic fluid glucose concentration: a rapid and simple method for the detection of intraamniotic infections in pre-term labor. Am J Obstet Gynaecol 1990; 163:968-974. 26. Hagenfeldt L, Hagenfeldt K: Individual free fatty acids in amniotic fluid and in plasma of pregnant women. B r J Obstet and Gynaecol 1976;83:383. 27. Biezenski JJ, Pomerace W, Goodman J: Studies on origin of amniotic fluid lipids. Am J Obstet Gynaecol 1968;143:479. 28. Soliman MK: Studies on physiological chemistry of allantoic and amniotic fluids of buffaloes at the various periods of pregnancy. Indian Vet J 1975; 52(2):106-112. 29. Klefin A, Suchanek E, Zobundzija M: Comparative study of allantoic adenosinetri phosphate activity and sodium and potassium concentration in bovine allantoic fluid. Veterinarski Archiv 1980;49(5):203-209. Cited in Vet Bull 50:7814:1020.
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JOURNAL OF EQUINE VETERINARY SCIENCE
COMPARATIVE BIOCHEMICAL INDICES OF FETAL FLUIDS IN NORMAL FOALING AND STRESSFUL DELIVERY IN INDIAN THOROUGHBRED MARES H.P.S. Kochhar, MVScl; P.S. Simran, MVSc2; A.S. Nanda, FRCVS, PhD1; Ripudaman Kaur, MVSc
SUMMARY
This study was conducted on 22 mares for establishing the vital biochemical parameters in the amniotic fluid and allantoic fluid of mares having normal parturition and difficult/stressful foaling (dystocia). Evaluation of the biochemical constituents in fetal fluids revealed that other than bilirubin, alkaline and acid phosphatase and lactate dehydrogenase, none of the biochemical parameter varied significantly amongst the two groups of normal foaling and dystocia suffering mares. The significant variation in the values of transaminases, estrases, creatine, urea nitrogen, glucose, sodium, potassium, calcium and phosphorus amongst the amniotic and allantoic fluids of each group (normal and dystocia) was observed. Variation in the biochemical indices could be attributed to fetal struggle and stress to the mare during dystocia. The allantoic and amniotic fluids vary widely in terms of biochemical constituents in each group.
INTRODUCTION
The volume of fetal fluids in mares, which is about 1020 liters towards term is regulated mainly by swallowing and excretion by the fetus. 1 The viscous amniotic fluid is derived from skin, mucous, amniotic epithelium fetal saAuthors' address:
1Department of Gynaecology & Obstetrics;
2Department of Surgery & Radiology, College of Veterinary Sciences, Punjab Agricultural University, Ludhiana-141 004 India. Acknowledgments: Authors wish to sincerely thank M/S Dasmesh Stud Farm, Sarai Naga, Faridkot, Punjab, India for providing the facilities.
206
liva and naso-pharyngeal secretions, while allantoic fluid primarily consists of excretion of fetal kidneys. 2 These fluids contain varying amounts of enzymes, phospholipids, sugars and ions, which are produced by the fetus. 3 The study of various biochemical constituents of fetal fluids could therefore serve as an index of fetal health even ante partum. Studies have been conducted on composition of fetal fluids in normal foaling 4 or in mid gestation, t The present study was planned with a view to establish the concentration of various enzymes and other vital constituents in fetal fluids of mares foaling normally or with assistance, i.e., dystocia. Efforts were made to correlate concentrations in both the groups, and establish variation if any in gestation length, fetal weight and placental weight in normal and difficult foaling mares.
MATERIALS AND METHODS
This study was conducted in the foaling season at an organized stud farm. In all 22 mares were taken into study, 11 having normal foaling and 11 requiring assistance up to moderate fetal mutations during foaling. The allantoic and amniotic fluids were collected by puncturing the respective sacs when presented in the birth canal, with the help of a sterile needle and syringe. Gestation length, parity, fetal sex and fetal weight, and placental weight were also recorded. The samples were transported on ice to the laboratory and were frozen at -20~ pending analysis within 3-5 days. The concentration of various biochemical constituents viz. glucose, 5 urea nitrogen, 6cholesterol, 7 aspartate aminotransferase 8 (AST), alanine aminotransferase 8 (ALT), alkaline JOURNAL OF EQUINE VETERINARY SCIENCE
phosphatase a (AKP), acid phosphatase a (ACP), acetylcholinesterase 9 (ACHE), butryl cholinesterase 10 (Butryl ChE), carboxylestrase 01 (Crb E), arylesterase 12 (ARE), lactate dehydrogenase 8 (LDH) and phosphorus la were determined by standard methods. Total proteins, albumin, bilirubin, creatinine were measured by standard kits. a Sodium (Na), potassium (K) and calcium (Ca) were estimated using Atomic Absorption Spectrophotometer. b Paired 't' test was employed for statistical analysis of the data. TM
Table 1. P r e g n a n c y d a t a of n o r m a l f o a l i n g a n d d y s t o c i a suffering m a r e s . Dystocia n = 11
Normal foaling n = 11
Gestation period (days)
332+7.79
323.87+7.9
Age of mares
9.4+3.61
11.13+4.83
3+1.18
4.37+2.86
Foal wt. (Kg)
53.5+7.09
50+7.07
Placenta wt. (Kg)
5.3+ 1.18
4.75+0.43
5 :6
6 :5
No. of mares
(yrs) Parity
RESULTS
All the foals delivered were live and various observations of normal and dystocia affected mares is tabulated in Table 1. The foal and placental weights, gestation length and age of mares were not statistically different between the two groups. The allantoic fluid was of amber color, clear and watery in consistency while the amniotic fluid was whitish, viscid and transopaque. The fetal fluids were free from blood, meconium or intermixing. The concentration of AKP, ACD, AST, ALT, ACHE, Butryl ChE, Carb E, Aryl E, urea nitrogen and glucose were significantly higher in amniotic than in allantoic fluid (p < 0.05) of mares both with normal as well as with assisted delivery. On the contrary concentration of LDH, globulin, bilirubin and creatinine were significantly higher (p < 0.05) in allantoic fluid than in amniotic fluid of both normal and dystocia suffering mares. The concentration of total proteins, albumin, immunoglobulin and cholesterol were similar in both types of fetal fluids. The concentration of AST, ALT butyl ChE, Carb E, Aryl E, total proteins, globulins, albumin, creatinine, glucose and cholesterol in allantoic fluid of dystocia affected mares were not significantly different than those in the normal mares. However, a significantly higher than normal (p < 0.05 & p < 0.001) concentration of AKP, LDH, bilirubin and BUN and lower concentration of ACP were observed in allantoic fluid of dystocia affected mares. The amniotic fluid concentration of bilirubin and AKP were significantly higher (p < 0.05) in mares with dystocia as compared to normal foaling ones.
DISCUSSION
The fetoplacental unit is a dynamic system with a constant exchange between fetal fluid compartments and maternal circulation. 2 A variation in the concentration of various biochemical components of allantoic and amniotic fluids could have a significant relation to the fetal health and may indicate certain fetal diseases, cystic fibrosis and aKits: Spans Diagnostics Pvt. Ltd. 173-B, New Industrial Estate, Udhana (Surat), India. bAtomic Absorption Spectrophotometer (Model AA6) Varian Techtran, Melbourne, Australia
Volume 17, Number 4, 1997
Sex ratio (Male : Female)
other conditions described in humans. 15 The alkaline and acid phosphatases are correlated with fetal maturity, fetal bone formation and kidney functioning, t In early pregnancy the AKP activity is mainly of fetal intestine origin. Meconium release or fetal defecation in distress contributes to its increased activity. Presumably AKP activity represents entry of desquamated intestinal mucosal cells 16 into fetal fluids. The concentration of AKP was significantly increased both in allantoic and amniotic fluid of mares suffering from difficulty at parturition as compared to their normal counterparts. The values in normally foaling mares corresponded with earlier studies. 1No reference for comparison of studies in dystocia affected mares are available for confirmation. The ALT and AST values are indicative of fetal liver functioning and muscular injuries if any. Their significantly different values for the amniotic and allantoic fluid are suggestive of the nature of fluid in each compartment, i.e., intestinal and tracheal secretions in amniotic fluid and fetal urine in allantoic fluid. 17 The concentration of esterases viz. ACHE, Butryl ChE, Carb E, Aryl E in amniotic and allantoic fluids of dystocia affected mares was significantly (p < 0.05) lower than normal. Although AChE is generally associated with neural tube defects or defects leading to leakage of CSF during fetal formation; however, it has rarely been observed in normal fetuses 1s,t9 too. Information on the source of these enzymes in fetal fluids is obscure. Perhaps stress due to enhanced fetal moments or struggle during dystocia might be a contributing factor to the concentration of esterases amongst the two groups. Lactate dehydrogenase is contributed by all types of cells including exfoliative cells. 1 Its rise has been reported with advance in fetal age and fetal death, a The rise in LDH, in the fetal fluids of dystocia suffering mares could be attributed to enhanced cellular exfoliation, escape of fetal intestine contents or due to extra fetal muscular movements during struggle to 207
Table 2 . B i o c h e m i c a l i n d i c e s in fetal f l u i d s of n o r m a l f o a l i n g a n d d y s t o c i a s u f f e r i n g m a r e s . Parameter
Units
AKpwwxxyYzz
IU/L
25.63+2.38
ACP wwxxyY
IU/L
1.91 +0.8
AST xwx
IU/L
ALT wx
lU/L
LDH wxxyY
I U/L
122.3_+14.5
AChE wxy
nmol thiol formed min-1 ml-1
14.94_+14.14
4.22+3.12
26.40+2.28
21.50_+22.72
Butryl ChE wwxx
nmol thiol formed min-1 ml-1
5.41 _+4.75
2.54_+2.06
13.49_+4.31
13.40_+7.26
Carboxyl Est wxx
nmol indophenol formed min-1 ml-1
11.25_+3.93
12.22_+3.77
25.00_+18.78
22.78_+9.36
nmol phenol formed min-1
0.09_+0.02
0.13_+0.07
0.69-+0.92
0.02_+0.004
UN wwy
mg/dL
27.75_+9.48
40.17_+8.16
41.7_+7.2
48.0_+9.0
Total protein x
gm/dL
1.21_+0.34
1.40_+0.31
0.91_+0.2
0.97_+0.37
Globulin wxx
grn/dL
1.02_+0.2
1.18_+0.14
0.78_+0.12
0.82_+0.2
Albumin
gm/dL
0.08_+0.3
0.13_+0.5
0.04_+0.06
0.05_+0.3
Total BilirubinWWxxyYzZ
mg/dL
1.2_+0.5
1.85_+0.23
0.21_+0.12
0.85_+0.32
Creatinine wWxX
mg/dL
68.7_+14.9
72.3_+8.8
19.38_+4.67
20.14_+4.77
Immunoglobulins
g/dL
0.87_+0.32
0.93_+0.56
0.63_+0.28
0.72_+0.34
Glucose wwxx
mg/dL
5.8_+1.19
6.2_+3.4
20.1_+5.4
21.6_+8.2
Cholesterol
mg/dL
8.8_+1.2
7.9_+3.4
7.2_+2.6
6.8_+3.1
Sodium wwxx
meq/L
71 _+23.8
88_+20.1
146_+4.8
149_+6.2
Potassium wwxx
meq/L
26.2_+10.8
28.4_+8.0
9.2_+1.8
10.4_+2.6
Calcium wwxx
meq/L
31.2_+9.8
34.3_+6.6
11.6_+2.8
12.2_+8.6
Phosphorus wwxx
meq/L
22.3_+8,7
26.6_+11.3
4.0_+1.8
4.4_+2.2
Aryl Est wxxzz
1) Normal Foaling:
Allantoic fluid vs Amniotic fluid w - differs significantly at p < 0,05 W - differs significantly at p < 0.001
Allantoic fluid Normal Dystocia 34.36+4.61
96.95+10.15
103.23_+13.11
1.3+0.4
6.59+1.51
8.44_+2.85
6.94_+3.38
7.48_+4.45
14.01 _+3.63
17.02_+10.91
9.70_+7.52
9.09_+7.99
24.78+13.94
16.92_+10.86
98.17_+ 18.2
112.8+21.4
178.0_+52.4
ml-1
3) Allantoic fluid: 4) Amniotic fluid:
2) Dystocia suffering mares: Allantoic vs Amniotic x - differs significantly at p < 0.05 X - differs significantly at p < 0.001
208
Amniotic fluid Normal Dystocia
Normal vs Dystocia suffering y - differs significantly at p < 0.05 Y - differs significantly at p < 0.001 Normal vs Dystocia z - differs significantly at p < 0.05 Z - differs significantly at p < 0.001
JOURNAL OF EQUINE VETERINARY SCIENCE
take birth. In dystocia affected animals the amniotic protein values were relatively lower than normal which is similar to that reported in bovines, a Concentration of bilirubin, a product of bile is indicative of liver health of animals. Enhanced biliary secretions may be the cause of increased bilirubin in both types of fetal fluids of dystocia suffering mares. In humans it has been reported that amniotic fluid bilirubin concentration becomes higher than in the maternal serum after 34 weeks of pregnancy probably due to decreased excretory ability of the maternal liver. 2~ Fetal urine contributes significantly to the allantoic fluid. 1 This may explain the rise of urea nitrogen and creatinine in stressfully foaling mares because of increased fetal micturition during violent efforts at the time of birth. Also in pregnancy creatinine moves from mother to fetal serum and then to fetal urine and to anmiotic fluid. 21 At times, however, creatinine may originate from fetal muscles and increased protein metabolism, a~ The glucose from maternal circulation is the main energy source for the fetus during normal pregnancy in ovines, aa although some of it is reduced to sorbitol in the placenta, a3 Our findings of lower glucose in the allantoic than amniotic fluid at foaling are in conjunction with those in bovines. 24 Increased glucose concentration in amniotic fluid has also been taken as an index of infectious state in pre-term labor in women, as Absence of any change from normal values in dystocia mares suggest it to be free from infections. It has been established that fetal renal excretion is the probable origin of free fatty acid in amniotic fluids of women. 26 The cholesterol represents one-third of total lipid profile in amniotic fluid but then it remains unchanged despite increase in quantity of amniotic fluid nearing term. 27 Immunologic activity of amniotic fluid remains poorly understood. 17 Allantoic fluid had much lower concentration of sodium than in amniotic fluid suggesting that the latter closely resembles the composition of plasma transudate and the flow of Na + from serum of mother to amniotic fluid is easy. 28'1 The high concentration of potassium in allantoic fluid is suggestive of increased fetal urine and metabolic activity of the fetus at tenn. Potassium ions differ in biological activity from sodium, as they cannot undergo passive transfer across cell membranes.29 High concentration of calcium in allantoic fluid is again attributed to fetal kidney status. The different biochemical constituents established in the above study are an effort to report the variation due to different foaling processes. Stress of dystocia, fetal movements in distress and forceful expulsive efforts by the mother may have contributed to variations in the values of some vital parameters. The endocrine studies may be correlated with such indices in an effort to bring forth the effect of stress of dystocia on biochemical constitution of fetal fluids in mares. Volume 17, Number 4, 1997
REFERENCES 1. Williams MA, Wallace Ss, Tyler JW, McCall CA, Gutierrez A, Spano JS; Biochemical characteristics of amniotic and allantoic fluid in late gestational mares. Theriogenology 1993;40:1251257. 2. Baetz AL, Hubert WT, Graham CK: Changes of biochemical constituents in Bovine fetal fluids with gestoterial age. Am J Vet. Res 1976;37:1047-1052. 3. Mohamed AR, Noakes DE: Enzyme activities in amniotic fluid and maternal blood in cattle before and after induced foetal death and abortion. Br Vet J 1985;141 (1):49-59. 4. Schott HC, Mannsman RA: Biochemical profiles of normal equine amniotic fluid at parturition. Equine Vet J 1988;5:52. 5. Frankel S, Reitman S, Sonnenwirth AC: In: Gradwohl's Clinical Laboratory Methods and Diagnosis. 7th edn. Vol. 1 St. Louis, USA: CV Mosby Company 1970. 6. Evans RT: Manual and automated methods of measuring urea based on a modification of its reactions with diacetyl monoxine and thiosemicarbazide. J Clin Pathol 1968;21:527. 7. Zak B: Simple and rapid micro technique for serum total cholesterol. Am J Cl in Pathol 1957;28:583. 8. Wooton IDP: Microanalysis in Medical Biochemistry. 4th edition. J&A Churchill Ltd. London UK, 1964. 9. Morari K, Usbiyama S, Satoh T, KugaT: Enzyme induction by repeated adminstration of tetrachlorvinophos in rats. Toxicol Appl Pharmacol 1976;37:377. 10. Vors G, Sachsso K: Red cell and plasma cholinestrase activities in micro samples of human and animal blood determined simultaneously by a modified acetylcholine/DTNB procedure. Toxicol Appl Pharmacol 1970;16:764. 11. Mendoza CE, Shields JB, Phillips WE J: Distribution of carboxylesterase activities in different tissues of albino rats. Comp Biochem Physiol 1971 ;40:841. 12. Mc Ilvain JE, Timoszyk J, Nakatsugawa T: Rat liver paraxonase (paraxona arylesterase). Pesticide Bioch and Physiol 1984;21:162. 13. Verlag H: Practical Clinical Biochemistry. 5th edn. William Hienemann Medical Books Ltd. London, 1980. 14. Snedecor GW, Cochran WG: Statistical methods 8th edition. Calcutta, India: Oxford and IBH Publishing Co. 1967. 15. Szabo M, Veress L, Teichmann F, Munich A, Huszka M, Papp Z; Amniotic fluid microviilar enzyme activity in fetal malformations. Clin Genet 1990;38:340-345. 16. Hahnemann N, Sorenson SA: Studies on alkaline phosphatase in amniotic fluid. Acta Obstet Gynaecol Scand 1974; 53:15-22. 17. Gleicher N: Immunological influence of amniotic fluid. Lancet 1980;1:541. 18. Barlow RD, Cuckle HS, Wald NJ: False positive gel acetylcholinestrase results in blood stained amniotic fluids. Br J Obstet Gynaecol 1982;89:821-829. 19. Goldfine C, Hadow J, Hudson G: Densitometry as an aid in amniotic fluid gel acetylcholinestrase analysis. Am J Obstet Gynaecol 1983; 145:317. 20. Benzie RJ, Doran TA, Harkins JL, Owen VMS, Parker CJ: Composition of amniotic fluid and maternal serum in pregnancy. Am J Obstet Gynaecol 1974;119:798. 21. Hodari AA, Mariona FG, Houliham RT: Creatinine transport in the maternal fetal complex. Obset Gynaecol 1973;41 : 47. 22. Alexander DP, Britton HG, Nixon DA: The metabolism of fructose and glucose by the sheep foetus: Studies on isolated perfused preparation with radioactivity labeled sugars. J Exp Med 1970;55:346-362. 23. Bdtton HG, Hugget ASG, Nixon DA: Carbohydrate metabolism in sheep placenta. Biochem BiophysActa 1967;136: 426-440. 24. Reeves JT, Daoud MD, Gentry M, Eastin C: Changes in
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urinary flow in bovine fetuses during late gestation: Comparison of amniotic and fetal body fluids. Am J Vet Res 1972;33(11 ):21592167. 25. Romero R, Jimenez C, Lohda AK, Neres J, Hanaoka S, Avila C, Callahan R, Mazoi M, Hobbins JC, Diamond MP: Amniotic fluid glucose concentration: a rapid and simple method for the detection of intraamniotic infections in pre-term labor. Am J Obstet Gynaecol 1990; 163:968-974. 26. Hagenfeldt L, Hagenfeldt K: Individual free fatty acids in amniotic fluid and in plasma of pregnant women. B r J Obstet and Gynaecol 1976;83:383. 27. Biezenski JJ, Pomerace W, Goodman J: Studies on origin of amniotic fluid lipids. Am J Obstet Gynaecol 1968;143:479. 28. Soliman MK: Studies on physiological chemistry of allantoic and amniotic fluids of buffaloes at the various periods of pregnancy. Indian Vet J 1975; 52(2):106-112. 29. Klefin A, Suchanek E, Zobundzija M: Comparative study of allantoic adenosinetri phosphate activity and sodium and potassium concentration in bovine allantoic fluid. Veterinarski Archiv 1980;49(5):203-209. Cited in Vet Bull 50:7814:1020.
Third Annual Cruise with Dr. John Madigan October 24-27, 1997 Dr. John Madigan Dipl American College Veterinary Internal Medicine University of California, Davis
Sail on Royal Caribbean's Viking Serenade to Ensenada, Baja, Mexico, from Long Beach, California. Six hours of continuing education credit will give you plenty of time to cruise the shops in Ensenada, Mexico. $349 pp plus port charges ($78.50) and meeting registration ($50), not including airfare. No registration fee for spouses or childdren. Low cost air fare available. 3rd & 4th passengers sail for $127.50 including port charges and taxes. Fares include stateroom, all meals, entertainment and most on-board activities. Topic: New Developments in Equine Medicine: From Research Findings to Practical Utilization. Contact: Ann Jones Journal of Equine Veterinary Science Phone (.909) 678-1889 r-ax Tor inrormadon (909) 678-1885. E-mail vetdata@ inland.net 210
JOURNAL OF EQUINE VETERINARY SCIENCE