Adlan M. Elhassan

Effects of estrogen and progesterone treatment on rat hippocampal NMDA receptors: relationship to Morris water maze performance.

Estrogen modulates NMDA receptors function in the brain. It increases both dendritic spine density and synapse number in the hippocampus, an effect that can be blocked by NMDA antagonist. In this study, we investigated the effect of 17β‐estradiol and progesterone treatment on NMDA receptors in ovariectomized rats. Two different doses were used for 10 weeks. Receptor autoradiography was done on brain sections using [3H] MK‐801 as a ligand. Our results showed a significant increase in [3H] MK‐801 binding in the dentate gyrus, CA3 and CA4 areas of the hippocampus of ovariectomized compared to sham operated rats. In addition, we observed similar changes in CA1. 17β‐estradiol treatment in both doses reduced the binding back to the normal level while progesterone treatment did not show any effect. Spatial reference memory was tested on Morris water maze task. Ovariectomy severely impaired spatial reference memory. Estradiol but not progesterone treatment significantly improved the memory performance of the ovariectomized rats. Low dose treatment showed better learning than high dose estrogen treatment. The decrease in the antagonist sites by estradiol treatment could result in an increase in the sensitivity of the hippocampus to the excitatory stimulation by glutamate system and hence the effect of estradiol on learning and memory. The changes of NMDA receptors in the hippocampus support the concept that estrogen‐enhancing effect on spatial reference memory could be through the enhancing of NMDA function.

Adjuvant-induced arthritis: IL-1 beta, IL-6 and TNF-alpha are up-regulated in the spinal cord.

Adjuvant-induced arthritis (AIA) is a widely used animal model of human rheumatoid arthritis (RA). We have previously shown that increased neuropeptide expression is observed in the spinal cord of AIA rats. To study the potential role of cytokines in the spinal cord of AIA, we wanted to determine whether there are changes of glial and cytokine expression (IL-1β, IL-6, TNF-α and IFN-γ) in the spinal cord of AIA rats. Our data indicated that macroglia and MHC class II immunostaining were enhanced, astrocytes expressing GFAP were increased in number and immunostaining intensity. Using in situ hybridization and immunohistochemical methods, both mRNA and protein levels of IL-1β, IL-6 and TNF-α were significantly increased in the spinal cord of arthritic rats. Increased cytokine expression was presented in the reactive astrocytes and microglia.

Suppression of autoimmune neuritis in IFN-γ receptor-deficient mice

1 T cells mediate demyelination in the peripheral nervous system (PNS). Infiltrating macrophages and T cells as well as cytokines like interferon (IFN)-g are intimately involved in causing pathogenic effects. To investigate the role of IFN-g in cell-mediated EAN, IFN-g receptor-deficient mutant (IFN-gR 2/2 ) C57BL/6 mice and corresponding wild-type mice were immu- nized with P0 peptide 180 -199, a purified component of peripheral nerve myelin, and Freunds complete adjuvant. IFN-gR 2/2 mice exhibited later onset of clin- ical disease. The disease was also less severe than in wild-type mice. Fewer IL-12-producing but more IL-4- producing cells were found in sciatic nerve sections from IFN-gR

Methionine-enkephalin in bone and joint tissues.

Methionine‐enkephalin (met‐enk), an endogenous opiate, mimics many of the effects of morphine by binding to opiate receptors, thereby eliciting similar cellular and behavioral effects. Using biochemical and immunohistochemical techniques, several peptides have been identified in bone and joint tissues. Here we report, for the first time, the presence as well as concentration of met‐enk in bone and joint tissues. Immunohistochemistry using electron and immunoflourescence microscopy showed cellular and neuronal distribution of met‐enk in bone and joint tissues. The concentration of met‐enk analyzed by high performance liquid chromatography electrochemical detection or radioimmunoassay was high in bone marrow, periosteum, ankle joint tissue, and cortical bone. Analysis by fast atom bombardment mass spectrometry suggested that the recovered fragment was met‐enk. Administration of met‐enk inhibits osteoblast cell growth in culture, which is reversible by naltrexone. In arthritic rats, the concentration of met‐enk was significantly decreased in ankle joints compared with controls, suggesting a role for met‐enk in the pathophysiology of adjuvant arthritis.

CD28–B7 costimulation: a critical role for initiation and development of experimental autoimmune neuritis in C57BL/6 mice

CD28 provides a critical costimulatory signal for antigen-specific T cell activation. Because CD28 is an important factor in the development of autoimmune diseases, we investigated its role in T cell-mediated experimental autoimmune neuritis (EAN), an animal model of Guillain-Barré syndrome in humans. CD28-deficient mutant (CD28-/-) C57BL/6 mice and corresponding wild-type mice were immunized with P0 peptide 180-199, a purified component of peripheral nerve myelin, and Freunds complete adjuvant. As a result, all wild-type mice developed severe EAN, in contrast, none of the CD28-/- mice manifested clinical signs of disease. Additionally, CD28-/- mice had fewer IL-12 producing cells in sciatic nerve sections and fewer IFN-gamma secreting splenic cells than wild-type mice on day 24 post immunization, i.e., at the peak of clinical EAN. At that time point, CD28-/- mice had milder infiltration of such inflammatory cells as macrophages, CD4+ T cells and monocytes into sciatic nerve tissues and less demyelination than wild-type mice. Moreover, the CD28-deficiency led to reduced production of specific anti-P0 peptide 180-199 antibodies compared with wild-type mice. Evidently, CD28 is required for interaction with B7 to regulate the activation of T and B cells that initiates development of EAN.

Loss of muscarinic M4 receptors in spinal cord of arthritic rats: implications for a role of M4 receptors in pain response

Changes in the levels of muscarinic M4 receptors in spinal cord of acute and chronic arthritic rats (animal models of pain) were studied by receptor autoradiography using muscarinic M4 receptor subtype selective ligand. Arthritis was induced in female Lewis rats by single intradermal injection of heat-killed Mycobacterium butyricum and sacrificed 12 days (acute group) and 30 days (chronic and control groups) after induction of arthritis. Our results demonstrate significant reduction in the level of M4 receptors in the spinal cord (Rexed laminae I-X) of acute and chronic arthritic rats compared to controls. These findings suggest that the muscarinic M4 receptor subtype may be involved in cholinergic mechanisms of analgesia.

Somatostatin immunoreactivity in bone and joint tissues

USING immunoelectron microscopy we have investigated the presence of somatostatin in normal bone and joint tissues. We observed somatostatin labeling in the myelinated nerve fibers of the periosteum, the bone marrow cells and in the mature bone matrix but only slightly in the synovial cells. Quantification of somatostatin in bone tissue by radioimmunoassay showed highest levels in bone marrow followed by periosteum and cortical bone. These findings suggest a role for somatostatin in bone and joint physiology.

Prolactin, growth hormone, and IGF-1 in ankles and plasma of adjuvant arthritic rats.

In this study we have investigated the levels of prolactin, growth hormone, and insulin-like growth factor-1 in plasma and in tissue extracts of ankle joints of rats with acute or chronic adjuvant arthritis using enzyme immunoassay (EIA) and radioimmunoassay (RIA). We found a stable content of prolactin in plasma of the different groups but a significantly increased concentration of growth hormone was observed in the plasma of the group with chronic arthritis. Moreover, an increased concentration of insulin-like growth factor-1 was noted in the plasma of the acute group. This evidently had returned to normal levels in the chronic group. In contrast, decreased concentrations of prolactin, growth hormone, and insulin-like growth factor-1 were found in tissue extracts of ankle joints of the group with chronic arthritis. The changes in the levels of these hormones in adjuvant arthritis might suggest that they play a role in the pathogenesis of the disease. Understanding the mechanism(s) of hormonal participation in adjuvant arthritis may open new treatment strategies for rheumatoid arthritis and other inflammatory disorders.

Intracerebroventricular administration of somatostatin prevents and attenuates adjuvant arthritis

The effects of somatostatin on the development of adjuvant arthritis induced by Mycobacterium butyricum were studied. Somatostatin was injected into the lateral cerebral ventricle every day for 14 days beginning on the first day of mycobacteria inoculation in the preventive group. In the treatment group, somatostatin was injected from day 17 until day 30 post-mycobacteria inoculation. Arthritis was evaluated by measuring ankle joint circumference and diameter as well as microscopic examination of ankle joint sections. Somatostatin profoundly inhibited the development of adjuvant arthritis and an anti-inflammatory action was observed in the treatment group. These results suggest that somatostatin has a central action that can prevent or attenuate symptoms associated with arthritis.

Effects of short- and long-term rat hind limb immobilization on spinal cord insulin-like growth factor-I and its receptor.

In this study we investigated changes in the spinal cord insulin-like growth factor-I peptide (IGF-I) and its receptors (IGF-IR) after hind limb immobilization for 5 days, 2, 4, and 8 weeks. Moreover, effects on IGF-I and nicotinic cholinergic receptors (nAChRs) in two types of skeletal muscle were also investigated. IGF-I levels were measured by radioimmunoassay (RIA) whereas IGF-IR and nAChRs were measured by quantitative receptor autoradiography. Spinal cord IGF-I levels decreased significantly after 5 days, 2 and 4 weeks of immobilization, whereas IGF-IR increased significantly after 4 and 8 weeks compared to controls. In skeletal muscles, nAChRs increased significantly after 5 days and 2 weeks in the soleus (SOL) and tibialis anterior (TIB) muscles, respectively, and continued up to 8 weeks in both muscles. IGF-I concentration decrease significantly after 4 and 8 weeks in the SOL and TIB muscles, respectively. Despite the normal levels of IGF-I in both muscles at the early time points (5 days and 2 weeks), low levels of IGF-I were observed concurrently in the spinal cord ipsilateral to the immobilized limb. Our findings suggest that the early decrease in the IGF-I level and the late upregulation in the IGF-IR in the spinal cord might represent a nervous system response to disuse.