Isolation, expansion, and characterization of mesenchymal stem cells from adult rat bone marrow

Mesenchymal stem cells [MSCs] have been studied and applied extensively because of their ability to self-renew and differentiate into various cell types. To isolate, increase the expansion rate and define rat bone marrow MSCs by immunohistochemical and electron microscopic studies. Twenty adult male albino rats were used and divided equally into two groups. The bone marrow was harvested and flushed out from the long bones of each group. In group 1, the cells were cultured using complete medium containing Dulbecco's modified Eagle's medium, 1% antibiotics, and 10% fetal bovine serum. In group 2, the cells were cultured in complete medium supplemented with 8% fetal bovine serum and 2% autologous rat serum. When the primary culture became nearly confluent, the adherent cells were subcultured. After 12 days from the primary culture, aliquots of the cells were prepared for Giemsa stain, transmission electron microscopy, and immunohistochemical staining for CD44, CD105, and CD34. The adherent cells in both groups were heterogeneous in appearance, and most of them were spindle or star-shaped with vesicular nuclei. Some cells were binucleated. The MSCs in group 2 reached confluency more rapidly than those in group 1. After passaging of group 2, the adherent cells became more homogeneously fibroblast-like in appearance. Immunostaining of MSCs revealed a positive reaction for CD44 and CD1 05 and a negative reaction for CD34. Ultrastructural examination revealed that the native MSCs appeared with many pseudopodia, the nucleus was eccentric, and the inner zone of the cytoplasm was rich in free ribosomes, many mitochondria, few rough endoplasmic reticulum, with obvious Golgi complex, and some lysosomes. Some MSCs showed no pseudopodia with central nucleus. Others appeared with two euchromatic elliptical nuclei and a nucleolus in one of them. MSCs can be easily purified, cultured, and expanded more rapidly using autologous rat serum

Immunological study of expression of CD1 in preimplantation and midgestation mouse embryo cells and placental trophoblasts and the role of decidual natural killer T cells at the feto-maternal interface

Expression of major histocompatibility complex molecules in the preimplantation embryo is important for control of development and immune recognition of the embryo by the mother. Four types of major histocompatibility complex class I molecules that have been identified are as follows: class Ia, class Ib, class Ic, and class Id. Expression of class Ic and class Id antigens in preimplantation embryos has not yet been fully evaluated. Cluster of differentiation 1 [CD1] molecule is a member of class Id antigen. Localization and regulation of CD1 expression remain unclear. The aim of this study was to detect expression of the CD1 molecule in a preimplantation mouse embryo at two-cell, four-cell, eight-cell, and blastocyst stages and in the embryo cells and placental trophoblasts at 10 days of gestation. Detection of invariant natural killer T [iNKT] cells in the decidua at the site of attachment of the placenta was also investigated. Mice of the C57BL/6 genetic background were used. Forty-eight female and 16 male mice of 10-12 weeks of age were divided into two groups. Group 1 included 24 female and eight male mice. Eight breeding colonies were constructed in which each cage contained one male and three female mice. The pregnant female mice were killed at 36, 48, 60, and 80 h of appearance of the vaginal plug to collect two-cell, four-cell, eight-cell, and blastocyst stages of embryos, respectively. The embryos were then processed for immunohistochemistry. Group 2 animals included 24 female and eight male mice. Eight breeding colonies were constructed in which each cage contained one male and three female mice. The pregnant female mice were killed at 10 days of gestation when embryo cells, placental cells, and decidual cells were isolated and processed for culturing and immunohistochemistry. This study showed no expression of CD1 molecules in any of the preimplantation mouse embryo stages till the blastocyst stage, but were detected on the cell surface and inside the cytoplasm of embryo cells and placental trophoblasts at 10 days of gestation. A high proportion of iNKT cells was also detected in the decidua at the site of attachment of the placenta. The CD1 molecule is not actively involved in the regulation of early development of embryo till the blastocyst stage, and the true biological role of CD1 in preimplantation development remains to be determined. However, it may play a role in the placenta and in embryo development during midgestation by preventing lysis of fetal and placental cells through an interaction with decidual iNKT cells at the feto-maternal interface

Effect of diet restriction on the survival of myenteric neurons in jejunum of aging male albino rat. histological and immunohistochemical study

Dysfunctions of the gastrointestinal system are more common complaints of the elderly. Enteric neurons are known to be vulnerable to age related cell death. So, the aim of this study was to study the effect of diet restriction on the histological changes in the myenteric neurons of jejunum of aged male albino rat. Thirty male albino rats of 6 months were used. Group I was the control group. Subgroup Ia included 5 rats of 6 months and subgroups Ib and Ic included 5 animals each and were continued to live up to the age of 12 and 18 months, respectively. Group II included 10 rats which were early fed the restricted diet [25 gm diet/rat! day] and subdivided into two subgroups five animals each. Subgroup ha were fed the restricted diet starting from 6 to 12 months. Subgroup IIb were fed the restricted diet starting from 6 to 18 months. Group III included 5 rats that were lately fed the restricted diet from 12 to 18 months. At time of sacrifice all the animals were anaesthetized, the jejunum was dissected out and specimens were processed for histological examination. Immunohistochemical expression of the pan-neuronal marker PGP 9.5 and neuronal nitric oxide synthase [nNOS] was investigated. As age advances in subgroup lb and Ic there was increased neuronal loss and obvious gaps in the myenteric ganglia with significant decrease in the number of PGP positive neurons. Animals of subgroup ha and llb showed that most of the myenteric neurons maintained their usual structural arrangement and expression of PGP. In group III rats showed neuronal loss and significant decrease in the PGP positive neurons. There was non significant decrease in the number of nNOS positive neurons in all groups. The results of the current study revealed that age related change involved mainly the cholinergic neurons and starting diet restriction early at 6 months age might slow or delay the age related damage of the rat myenteric neurons

interrelation of migraine, vestibular dysfunction and hearing impairment

Migraine is a relatively common neurological disorder. Neuro-otological symptoms were common in patients with migraines. The aim of this work is to identify the frequency of neuro-otological affection in migraine. The study includes forty-one patients with migraine, and 20 healthy volunteers. Each subject was submitted to full neurological, medical and audiological history and examination. Puretone audiometry, electronystagmography [ENG] tests, auditory brain stem response [ABR] and event related potentials [ERPs]. Dizziness was recorded in 56.1%, rotational vertigo in 31.7% and positional vertigo was occurred in 19.5%. Sense of imbalance, tinnitus and phonophobia were reported in 12.2%, 19.5%, and 29.3%, respectively. Abnormalities in at least one of ENG tests were found in 75.6% of patients that include abnormal results of positioning and positional in 34.1%, post head shaking test in 26.8%, saccade test in 19.5%, optokinetic test in 19.5%, pursuit test in 14.6%, unilateral caloric weakness test in 9.8% and spontaneous nystagmus test in 2.4%. Sensorineural hearing impairment was reported in 21.9%. Significant prolongation of mean interpeak latency of I-III of right ear at 90 dBHL with low repetition rate, III-V at 90 dBHL with high repetition rate and absolute latency of right wave V at 90 dBHL repetition rate compared with controls. No significant differences between patients and control in ERPs. Vestibular dysfunctions were frequently associated with common migraine also hearing impairment was reported in appreciable numbers of cases. Both central and peripheral vestibular affection were recorded in between attacks

effect of patients positioning on oxygen satration in the acute phase of ischemic stroke

Body position is known to influence respiratory function in normal subjects and those with respiratory pathology. Its effect on respiratory function after stroke has received little attention. It's generally thought that stroke morbidity is increased by inappropriate position during early recovery phase. So the present study was designed to identify changes in arterial oxygen saturation [SaO[2]] associated with different four positions of the patient in order to detect proper position which maintain the optimum level of SaO[2]. Quasiexperiment research design was applied in this study .This study was conducted in stroke intensive care unit of Neurological Department of Assuit University Hospitals. The subjects of this study consist of 60 patients with acute ischemic stroke within 48 hours following mild to moderate and sever stroke. Three tools were included in the study, tool of sociodemographic data, tool of the positions and tool of the Scandinavian stroke scale. The result of the present study revealed that there was statistical significant difference between SaO[2] and blood pressure of the patients before and after the positions. It was found also that total mean of SaO[2] during right side, supine and semi-setting position was nearly equal and there was statistical significant difference between left position and 3 other positions. So the study recommended the importance of changing patient's positions which improve ventilation and blood pressure. The importance of the use of positions tested [Rt.side, supine and semi-setting position] in the clinical practice to maintain SaO[2] in patients of ischemic stroke

Diagnostic value of PRO C global test in patients with deep vein thrombosis

This study was carried out to evaluate pro C Global [PCG] test in clinical routine with special regard to its sensitivity and specificity for factor V [FV] leiden as well as the deficiency of protein C [PC] and protein S [PS]. 70 adult patients with documented diagnosis of deep vein thrombosis [DVT] by Doppler ultrasonography were chosen from those were attending the emergency unit of Alexandria main university hospital and Alexandria armed forces hospital in addition to 30 age and sex matched healthy controls were evaluated for PCG test in relation to gold standard tests i.e. PC activity, PS activity and activated protein C resistance [APCR]. Also determination of lupus anticoagulants [LA] was done for all subjects under study. The sensitivity and specificity of PCG test were [100% and 100%] for FV leiden, [87.5% and 82.3%] for PC and [80% and 78.5%] for PS. The negative predictive value was [100%, 98.1% and 98.1%] for FV leiden, PC and PS respectively. The positive predictive value was [100%, 38.9% and 22.2%] for FV leiden, PC and PS respectively. Also, the diagnostic accuracy was [100%, 82.9% and 78.6%] for FV leiden, PC and PS respectively. The results of LA were negative in all patients and controls. However, the normalized ratio [NR] of PCG test was decreased in [14.8%] of patients group without any detectable defect in PC system and their results were significantly lower than control group [P=0.000]. On the other side, the results were considerably higher than those for the patients with a proven defect in PC system. Pro C Global test is sensitive, specific, less time consuming and can be performed on a routine base. Because of the high negative predictive value, we recommend the use of Pro C Global test in the screening of thrombophilic patients and further determination of F V leiden, PC activity and PS activity is only indicated in case of abnormal Pro C Global results