Figure 4 - uploaded by Iram Iqbal
Content may be subject to copyright.
Transverse section of 48 hours embryo of experimental group three (G3), under light microscope-Open neural tube.
Source publication
To describe the effect of high dose progesterone (HDP) alone, or in combination with folic acid (FA), on occurrence of neural tube defects (NTDs) in chick embryo. MATERIAL and
60 Fertile, specific eggs of Fyoumi species of chick were selected at zero hr of incubation. They were incubated at 37.5 °C and 75% relative humidity until the embryos reache...
Contexts in source publication
Context 1
... of defective regions of the embryos directly under dissecting microscope and as whole mount and transverse section under light microscope revealed that the neural folds usually elevated normally, but convergence often failed to occur (Figure 3, 6). In many of the embryos with neural tube defects, the elevated neural folds actually diverged, flaring laterally (Figure 4, 5). The formation of neural tube defects in embryos treated with HDP were principally due to a failure of the elevated neural folds to converge toward the dorsal midline. Fusion occasionally failed to occur at various levels along the length of the spinal cord, but much more frequently fusion was inhibited only in the area of the posterior neuropore (Figure 3, ...
Context 2
... of defective regions of the embryos directly under dissecting microscope and as whole mount and transverse section under light microscope revealed that the neural folds usually elevated normally, but convergence often failed to occur (Figure 3, 6). In many of the embryos with neural tube defects, the elevated neural folds actually diverged, flaring laterally (Figure 4, 5). The formation of neural tube defects in embryos treated with HDP were principally due to a failure of the elevated neural folds to converge toward the dorsal midline. ...
Similar publications
Neural tube defects (NTDs) are the most common of the severe malformations of the brain and spinal cord. Increased maternal intake of folic acid (FA) during the periconceptional period is known to reduce NTD risk. Data from 1046 NTD cases in South Carolina were gathered over 20 years of surveillance. It was possible to determine maternal periconcep...
Citations
... Such alterations potentially lead to neurobehavioural changes in the adult offspring of Wistar rats [6]. Folic acid supplementation decreased the incidence of the defects neural tube caused by certain drugs as progesterone, but did not obviate them [7]. The toxicity with LD50 values of folic acid by the i.p. route in different strains of mice showed convulsions, ataxia and weakness. ...
The present study was conducted to evaluate the effect of folic acid (FA) administration in early pregnancy on spinal cord in mice fetuses. Virgin female albino mice, 8-10 weeks old were were obtained from the animal breeding house of veterinary medicine collage, Omar Al mukhtar University, EL Beida ,Libya. The animals mated 1 male: 2 female and next morning examined to confirmed successful mating by vaginal smear. Appearance of vaginal plug was considered a day zero of pregnancy. The pregnant females were divided into equal three groups. GI : Control group received orally distilled water, G II (Normal dose group) : given orally folic acid at dose level 80ug /kg bw for a week and G III(High dose group): given orally folic acid at dose level 160ug/kg bw for a week. Oral administration of folic acid was achieved by oral gavage from the 7 th day until the 14 th day of gestation then the pregnant females were observed till 18 th day of gestation (per natal). On 18 th day of gestation the contents of the uterus from both control and treated mice were examined with microscope for gross abnormalities and external morphological defects in the brain or spinal cord. Transverse sections of spinal cord at cervical, thoracic and lumbar levels were prepared and stained with haematoxylin and eosin (H&E) for histological study. Neither clinical signs nor abnormalities in behavior and external features were observed in pregnant mice treated with normal or high dose of folic acid. Also, no mortality was recorded in control and folic acid treated groups during experimental period. Microscopic examination of the embryos of mice treated with normal dose of folic acid revealed no obvious defects in the brain or spinal cord compared to control group. No outstanding differences in the mean number and body weight of fetuses between control and normal dose of folic acid treated group. While, high dose of folic acid induced marked decrease in the number and size of fetuses. In histological study administration of normal dose of folic acid revealed normal architecture of spinal cord regions with distinct grey and white matter, normal pattern of cellular density, nuclear shape and arrangement with slight increase in the extracellular space ,intact lining ependymal cells of central canal and normal widened anterior median fissure compared to the corresponding spinal cord regions of the control. While, administration high dose of folic acid in early pregnancy had adverse defects on mice fetuses. This study suggested that administration of normal dose of folic acid for a week in early pregnancy had no noticeable effect on spinal cord of mice fetuses .
... It is attributed to high doses of progesterone used to avoid fetal loss in these clinical scenarios. Iqbal et al [8] studied the effect of high dose progesterone on the development of neural tube using chick embryo model. They found NTD in 75% of the eggs treated with high dose progesterone while none of control group embryos developed this malformation. ...
Pre-conceptional supplementation of folic acid is well known to reduce the incidence of spina bifida. But Athena is frequently perplexed to see this anomaly occurring despite folate supplementation. In a largest Canadian study, De Wal et al [1] screened 1.9 million live births to study the effect of mass fortification. On comparing pre and post fortification periods they noted only 46% reduction in the incidence of neural tube defects (NTD). What happens to the remaining 54%? Why do they defy “folic acid” logics? Recently several publications have shed more light on this intriguing question.
AIm: To describe the effect of high dose progesterone (HDP) alone, or in combination with folic acid (FA), on occurrence of neural tube defects (NTDs) in chick embryo. mAterIAl and methOds: 60 Fertile, specific eggs of Fyoumi species of chick were selected at zero hr of incubation. They were incubated at 37.5 ºC and 75% relative humidity until the embryos reached stage eight of development. At this stage the eggs were divided into four groups consisting of 15 eggs/group. The 1st group was incubated without any operation. The 2nd group was injected with physiological saline. The 3rd and 4th groups were injected with HDP (20x physiologic dose of progesterone) and HDP with supplement of 5 micrograms/embryo of FA, respectively. After 48 hrs of incubation, all embryos were reviewed for the presence of NTDs under light microscopy. results: None of the eggs in the control, and saline injection groups showed NTDs, whereas 75 % (9/12) of the embryos in the 3rd group, and 58.3 % (7/12) of the chick embryos in 4th group showed NTDs. COnClusIOn: Exogenous progesterone at levels twenty times above its physiologic range in chick embryos causes NTDs. FA supplementation decreases the frequency of NTDs but does not abolish them. ÖZ AmAÇ: Yalnızca Progesteron ve progesterona ilave olarak Folik asit ile ile kombine edilmiş uygulamanın civciv embiryosunda nöral tüp defekti oluşumu üzerine olan etkisi. yÖntem ve GereÇ: Çalışmada, inkübasyonun 0. saatinde 60 fertil Fyoumi türü civciv embiryosu kullanıldı.Tüm embiryolar gelişimlerinin 8. evresine kadar, 37,5 ºC ve 75 % nemli ortamda inkübasyona tabi tutuldu. Bu evrede embiryolar 15'erli 4 gruba ayrıldı. Birinci grup herhangi bir işleme maruz bırakılmaksızın inkübasyona tabi tutuldu. İkinci gruba serum fizyolojik injeksiyonu yapıldı. Üçüncü gruba yüksek doz progesteron (fizyolojik dozun 20 katı), dördüncü gruba yüksek doz progesterona ek olarak 5 mikrogram folik asit injeksiyonu yapıldı. Bu işlemlerden sonraki 48 saat boyunca inkübe edilen embriyolar ışık mikroskobunda nöral tüp defekti gelişimi açısından incelemeye alındı. BulGulAr: Kontrol grubunda ve serum fizyolojik injeksiyonu yapılan grupta nöral tüp defekti saptanmazken, 3. grupta % 75 oranında (9/12), 4. grupta ise % 58,3 (7/12) oranında nöral tüp defekti geliştiği görüldü. sOnuÇ: Fizyolojik dozun 20 katında uygulanan progesteron civciv embriyolarında nöral tüp defekti oluşumuna neden olmaktadır. Folik asit desteği nöral tüp defekti oluşum sıklığını azaltsa da defektin görülmesini tam olarak ortadan kaldıramamaktadır.
Cisplatin is an effective antineoplastic drug. It has been used to treat ovarian, testicular, and bladder carcinoma. However, a serious disadvantage of this drug is the induction of various severe side effects in kidneys, bone marrow and liver. Cisplatin if administrated during pregnancy may cause many malformations including ventriculomegaly, microphthalmous and polyhydraminos. Neural tube defects are a multifactorial disorder, arising from a complex combination of genetic and environmental interactions, and while are only now beginning to understand the etiologies of neural tube defects, significant advances have been made in their prevention and treatment by taking folic acid (FA) during child-bearing years . Based on animal studies, epidemiologic studies and intervention trials, maternal folic acid is known to be protective for neural tube defects, primarily spinabifida and anencephalus.
This study was aimed to assess the prenatal and postnatal effect of cisplatin on spinal cord, dorsal root ganglion &sciatic nerve and to investigate the neuro-protective effect of folic acid against prenatal & postnatal cisplatin neurotoxicity.
Sixty adult female albino rats were used in this study. Animals were placed in quarantine for one week prior to breeding and housed in separate plastic cages in a controlled environment with free access to water and balanced diet. After acclimatization to laboratory conditions, two females were kept overnight with one male to allow mating; the presence of sperms in vaginal smear in the next morning indicates the day zero of pregnancy.
Two drugs were used in this study:-The cisplatin in the form of a vial contains 50 mg liquid cisplatin. . Every rat was injected intraperitoneally by 1 mg cisplatin /kg once every other day and this was equivalent to the human low therapeutic dose. The folic acid is in the form of 800 microgram tablets powdered and dissolved in 10 ml of distilled water, each rat was given 5ml of the solution once daily through a gastric tube.
Sixty pregnant rats were divided into two equal groups:
I- Prenatal group: contained thirty pregnant rats and was subdivided into three equal subgroups.
a. Prenatal control subgroup: received saline from 1st day of pregnancy till the time of sacrifice at 18th day& 20th day of pregnancy.
b. Prenatal cisplatin subgroup: were injected intraperitoneally with the therapeutic dose (l mg/Kg) of Cisplatin on alternate days from 1st. day of pregnancy till the time of sacrifice at 18th day& 20th day of pregnancy.
c. Prenatal cisplatin & folic acid subgroup: were injected intraperitoneally with (I mg/Kg) of Cisplatin (every other day) Plus folic acid in a dose of (400microgram) daily by the nasogastric tube from 1st day of pregnancy till the time of sacrifice at 18th day& 20th day of pregnancy.
The fetuses were obtained by caesarian section then the specimens of the spinal cord, dorsal root ganglion and sciatic nerve were obtained as by dissection then the specimens were then prepared for light and electron microscopic examination
II- Postnatal group: contained thirty pregnant rats and was subdivided into three equal subgroups.
a- Postnatal control subgroup: The pregnant rats were allowed to complete pregnancy. Their pups received saline from 1st day of delivery till being sacrificed on the 7th Day,14th Day & 21st day postnatally
b- Postnatal cisplatin subgroup: The pregnant rats were allowed to complete pregnancy. Their pups were injected intraperitoneally with the therapeutic dose of Cisplatin (l mg/Kg) on alternate days from 1st day of delivery till being sacrificed on the 7th Day, 14th Day & 21st day postnatally.
c- Postnatal cisplatin & folic acid subgroup: The pregnant rats were allowed to complete pregnancy. Their pups were injected intraperitoneally with the therapeutic dose of Cisplatin(I mg/Kg) every other day plus folic acid in a dose of (400microgram) daily given to the pups through a gastric tube from 1st day of delivery till being sacrificed on the7th Day,14th Day & 21st day postnatally .
The pups were anaesthetized by ether inhalation and the specimens were obtained by dissection. The specimens of the spinal cord, dorsal root ganglion and sciatic nerve were prepared for light and electron microscopic examination
In the present study, cisplatin has a toxic effect on the developing spinal cord. This toxicity is represented as degeneration in the grey and white matter. The shape of the spinal cord was severely affected and this effect ranges from disfigurement of the grey matter in fetuses aged 18 days to shrinkage of the spinal cord in pubs aged 14 days to end in complete degeneration of the cord which is replaced by large vacuoles and spots of hemorrhage in rats aged 21 days. Ultrastructure examination confirms this toxicity as the sections of the spinal cord show distorted neurons with eccentric piknotic nuclei, these nuclei consists mainly of heterochromatin and surrounded by irregular nuclear membrane, the cytoplasm is full of vacuoles and dilated rough endoplasmic reticulum and swollen mitochondria. In the current study, the folic acid can alter the cisplatin toxicity of the spinal cord as the specimens received folic acid during treatment with cisplatin show signs of improvement as, the vaculations in the grey and white matter decreased and the size of the spinal cord increased and its outlines are regular. The ultrastructure confirms that as, the amount of euchromatin increased in the nuclei, these nuclei have regular nuclear membrane. The cytoplasm of theses neurons is full of healthy mitochondria but some of them are swollen. The myelinated axons have no vacuoles in the myelin sheath or shrinkage of the axoplasm.
Regarding the dorsal root ganglia, in this study, the cisplatin has a toxic effect on the dorsal root ganglia. This toxicity appears in a mild form in the prenatal groups which show that the ganglia have dark neurons with small nuclei; there are wide spaces separate the neurons and the nerve fibers. Then the toxicity became sever on the older ages as ganglia of rats aged fourteen and twenty one days show that some neurons are piknotic with dark nuclei and others are degenerated with chromatolytic nuclei and the neuroglia are accumulated on one side as residual bodies the nerve fibers show large vacuoles. The ultrastructure of the ganglia confirmed the cisplatin toxic effect as the neurons are shrunken with pale nucleus and irregular nuclear membrane, the cytoplasm has large vacuoles and the mitochondria are the most subjected organ for the toxicity as these mitochondria are swollen and vacuolated. In this study, the folic acid has a minor protective role on dorsal root ganglia against cisplatin toxicity, as the specimens of the dorsal root ganglia that received folic acid during cisplatin therapy show that some neurons have large pale nucleus and prominent nucleolus. The ultrastructure of the dorsal root ganglia confirmed that the groups received folic acid show little improvement as the nuclear membrane of the neurons in these groups is more regular than that of the cisplatin group but the cytoplasm has many vacuoles these vacuoles decreased in older ages.
Regarding the sciatic nerve, the cisplatin has neurotoxic effects on the sciatic nerve; these toxic effects appear in the prenatal groups as multiple vacuoles and spots of hemorrhage in the nerve fibers. The signs of toxicity also appear in the postnatal groups that show swollen and degenerated axons with few Schwann cells and fibrosis in the nerve fibers with excess fibrous tissue in the perineurium. The ultrastructure confirms the cisplatin neurotoxicity as the sciatic nerve of the prenatal groups treated by cisplatin show unmyelinated fibers with vacuolated axoplasm, the axoplasm of these fibers contains few mitochondria. In this study the postnatal groups show shrinkage of axoplasm of the myelinated axons with few mitochondria and neurofibrils and vacuoles in the myelin sheath.
In this study the folic acid cannot protect the sciatic nerve against cisplatin toxicity as the prenatal groups received folic acid during cisplatin therapy show spaces in the nerve fibers. The postnatal groups show multiple vacuoles in the nerve fibers with areas of fibrosis some nerve fibers are degenerated. The ultrastructure confirmed that the folic acid cannot protect sciatic nerve against toxicity as the groups treated with folic acid during cisplatin therapy show shrinkage of the axoplasm with few mitochondria, neurofibrils and vacuoles in the axoplasm the surrounding myelin sheath is thin.
Conclusion:
Cisplatin has neurotoxic effects on the developing spinal cord, dorsal root ganglia and sciatic nerve. Folic acid can protect the spinal cord and dorsal root ganglia against these toxicities but cannot protect the toxicity of the sciatic nerve. So the pregnant female should be supplied by adequate folic acid doses during cisplatin therapy.
Recommendation:
We recommend further studies concerning other drugs that may prevent cisplatin neurotoxicity.
Although the prevalence of congenital malformations is high in China, little information is available on the Hainan Province.
Data used in this study were collected from 2000 to 2010 by monitoring newborns (including both live births and stillbirths) in the Hainan Province. Comparison analyses were conducted by performing χ(2) tests.
The data showed an increasing temporal trend of the birth defect prevalence in the Hainan Province; however, the rate of birth defects changed over time.
The Hainan Province is confronted with the challenge to reduce the prevalence of birth defects. Interventions should focus on the rural areas where a higher prevalence of birth defects is observed. Birth Defects Research (Part A) 97:750-754, 2013. © 2013 Wiley Periodicals, Inc.
