Ildiko Szalayova

Transplanted bone marrow generates new neurons in human brains.

Adult bone marrow stem cells seem to differentiate into muscle, skin, liver, lung, and neuronal cells in rodents and have been shown to regenerate myocardium, hepatocytes, and skin and gastrointestinal epithelium in humans. Because we have demonstrated previously that transplanted bone marrow cells can enter the brain of mice and differentiate into neurons there, we decided to examine postmortem brain samples from females who had received bone marrow transplants from male donors. The underlying diseases of the patients were lymphocytic leukemia and genetic deficiency of the immune system, and they survived between 1 and 9 months after transplant. We used a combination of immunocytochemistry (utilizing neuron-specific antibodies) and fluorescent in situ hybridization histochemistry to search for Y chromosome-positive cells. In all four patients studied we found cells containing Y chromosomes in several brain regions. Most of them were nonneuronal (endothelial cells and cells in the white matter), but neurons were certainly labeled, especially in the hippocampus and cerebral cortex. The youngest patient (2 years old), who also lived the longest time after transplantation, had the greatest number of donor-derived neurons (7 in 10,000). The distribution of the labeled cells was not homogeneous. There were clusters of Y-positive cells, suggesting that single progenitor cells underwent clonal expansion and differentiation. We conclude that adult human bone marrow cells can enter the brain and generate neurons just as rodent cells do. Perhaps this phenomenon could be exploited to prevent the development or progression of neurodegenerative diseases or to repair tissue damaged by infarction or trauma.

CD45‐Positive Blood Cells Give Rise to Uterine Epithelial Cells in Mice

The uterine endometrium is composed of epithelial and stromal cells, which undergo extensive degeneration and regeneration in every estrous cycle, and dramatic changes occur during pregnancy. The high turnover of cells requires a correspondingly high level of cell division by progenitor cells in the uterus, but the character and source of these cells remain obscure. In the present study, using a novel transgenic mouse, we showed that CD45‐positive hematopoietic progenitor cells colonize the uterine epithelium and that in pregnancy more than 80% of the epithelium can derive from these cells. Since we also found green fluorescent protein (GFP)‐positive uterine endothelial cells in long‐term GFP bone marrow‐transplanted mice, we conclude that circulating CD45+ cells play an important role in regenerating the uterine epithelium.

The combination of granulocyte colony-stimulating factor and stem cell factor significantly increases the number of bone marrow–derived endothelial cells in brains of mice following cerebral ischemia

Granulocyte colony-stimulating factor (G-CSF) induces proliferation of bone marrow-derived cells. G-CSF is neuroprotective after experimental brain injury, but the mechanisms involved remain unclear. Stem cell factor (SCF) is a cytokine important for the survival and differentiation of hematopoietic stem cells. Its receptor (c-kit or CD117) is present in some endothelial cells. We aimed to determine whether the combination of G-CSF/SCF induces angiogenesis in the central nervous system by promoting entry of endothelial precursors into the injured brain and causing them to proliferate there. We induced permanent middle cerebral artery occlusion in female mice that previously underwent sex-mismatched bone marrow transplantation from enhanced green fluorescent protein (EGFP)-expressing mice. G-CSF/SCF treatment reduced infarct volumes by more than 50% and resulted in a 1.5-fold increase in vessel formation in mice with stroke, a large percentage of which contain endothelial cells of bone marrow origin. Most cells entering the brain maintained their bone marrow identity and did not transdifferentiate into neural cells. G-CSF/SCF treatment also led to a 2-fold increase in the number of newborn cells in the ischemic hemisphere. These findings suggest that G-CSF/SCF treatment might help recovery through induction of bone marrow-derived angiogenesis, thus improving neuronal survival and functional outcome.

Transforming growth factor α induces angiogenesis and neurogenesis following stroke

The cytokine transforming growth factor alpha (TGF alpha) has proangiogenic and proneurogenic effects and can potentially reduce infarct volumes. Therefore, we administered TGF alpha or vehicle directly into the area surrounding the infarct in female mice that received gender-mismatched bone marrow transplants from green fluorescent protein (GFP)-expressing males prior to undergoing permanent middle cerebral artery occlusion. Newborn cells were tracked with bromodeoxyuridine (BrdU) labeling and immunohistochemistry at 90 days after stroke onset. We also studied the ingress of bone marrow-derived cells into the ischemic brain to determine whether such cells contribute to angiogenesis or neurogenesis. Infarct volumes were measured at 90 days poststroke. The results show that TGF alpha led to significant increments in the number of newborn neurons and glia in the ischemic hemisphere. TGF alpha also led to significant increments in the number of bone marrow-derived cells entering into the ischemic hemisphere. Most of these cells did not label with BrdU and represented endothelial cells that incorporated into blood vessels in the infarct border zone. Our results also show that infarct size was significantly reduced in animals treated with TGF alpha compared with controls. These results suggest that TGF alpha can induce angiogenesis, neurogenesis and neuroprotection after stroke. At least part of the pro-angiogenic effect appears to be secondary to the incorporation of bone marrow-derived endothelial cells into blood vessels in the infarct border zone.

Reversal of Sjögren’s-like syndrome in non-obese diabetic mice

Background: Non-obese diabetic (NOD) mice exhibit autoimmune diabetes and Sjögren’s-like syndrome. Objective: To test whether a treatment that reverses end-stage diabetes in the NOD mouse would affect their Sjögren’s-like syndrome. Methods: NOD mice have a proteasome defect. Improperly selected naive T cells escape, but can be killed by reintroducing major histocompatibility complex class I self-peptides on matched normal splenocytes. The proteasome defect also impairs nuclear factor kB, a transcription factor in pathogenic memory T cells, increasing their susceptibility to tumour necrosis factor-induced apoptosis stimulated through complete Freund’s adjuvant (CFA). The impact of this two-limb therapy (injections of matched normal splenocytes and CFA) on the autoimmune salivary gland disease of the NOD mice was studied. Results: All NOD mice receiving the above treatment had a complete recovery of salivary flow and were protected from diabetes. Restoration of salivary flow could be the result of a combination of rescue and regeneration of the gland, as confirmed by immunohistochemical analysis. All untreated NOD mice showed a continuous decline in salivary flow, followed by hyperglycaemia and death. Conclusion: This study establishes that a brief intervention in NOD mice with Sjögren’s-like syndrome can reverse salivary gland dysfunction.

Of splice and men: what does the distribution of IKAP mRNA in the rat tell us about the pathogenesis of familial dysautonomia?

Familial dysautonomia (FD) is the best-known and most common member of a group of congenital sensory/autonomic neuropathies characterized by widespread sensory and variable autonomic dysfunction. As opposed to the sensory/motor neuropathies, little is known about the causes of neuronal dysfunction and loss in the sensory/autonomic neuropathies. FD involves progressive neuronal degeneration, has a broad impact on the operation of many of the bodys systems, and leads to a markedly reduced quality of life and premature death. In 2001, we identified two mutations in the IKBKAP gene that result in FD. IKBKAP encodes IKAP, a member of the putative human holo-Elongator complex, which may facilitate transcription by RNA polymerase II. Whether or not the Elongator plays this role is moot. The FD mutation found on >99.5% of FD chromosomes does not cause complete loss of function. Instead, it results in a tissue-specific decrease in splicing efficiency of the IKBKAP transcript; cells from patients retain some capacity to produce normal mRNA and protein. To better understand the relationship between the genotype of FD patients and their phenotype, we have used in situ hybridization histochemistry to map the IKAP mRNA in sections of whole rat embryos. The mRNA is widely distributed. Highest levels are in the nervous system, but substantial amounts are also present in peripheral organs.

Neuronal M3 muscarinic acetylcholine receptors are essential for somatotroph proliferation and normal somatic growth

The molecular pathways that promote the proliferation and maintenance of pituitary somatotrophs and other cell types of the anterior pituitary gland are not well understood at present. However, such knowledge is likely to lead to the development of novel drugs useful for the treatment of various human growth disorders. Although muscarinic cholinergic pathways have been implicated in regulating somatotroph function, the physiological relevance of this effect and the localization and nature of the receptor subtypes involved in this activity remain unclear. We report the surprising observation that mutant mice that selectively lack the M3 muscarinic acetylcholine receptor subtype in the brain (neurons and glial cells; Br-M3-KO mice) showed a dwarf phenotype associated with a pronounced hypoplasia of the anterior pituitary gland and a marked decrease in pituitary and serum growth hormone (GH) and prolactin. Remarkably, treatment of Br-M3-KO mice with CJC-1295, a synthetic GH-releasing hormone (GHRH) analog, rescued the growth deficit displayed by Br-M3-KO mice by restoring normal pituitary size and normal serum GH and IGF-1 levels. These findings, together with results from M3 receptor/GHRH colocalization studies and hypothalamic hormone measurements, support a model in which central (hypothalamic) M3 receptors are required for the proper function of hypothalamic GHRH neurons. Our data reveal an unexpected and critical role for central M3 receptors in regulating longitudinal growth by promoting the proliferation of pituitary somatotroph cells.

Localization of S100A8 and S100A9 expressing neutrophils to spinal cord during peripheral tissue inflammation.

&NA; Investigation of hyperalgesia at the spinal transcriptome level indicated that carrageenan‐induced inflammation of rat hind paws leads to a rapid but sustained increase in S100A8 and S100A9 expression, two genes implicated in the pathology of numerous inflammatory diseases including rheumatoid arthritis and gout. In situ hybridization revealed that the elevation occurred in neutrophils that migrate to the spinal cord vasculature during peripheral inflammation, not in spinal neurons or glial cells. Immunohistochemical analysis suggests, but does not prove, that these neutrophils abundantly release S100A8 and S100A9. Consistent with this, we detected an increase in ICAM and VCAM, both indicators of endothelial activation, a known trigger for secretion of S100A8 and S100A9. Migration of S100A8‐ and S100A9‐expressing neutrophils to spinal cord is selective, since MCP‐1‐ and CD68‐expressing leukocytes do not increase in spinal cord vasculature during hind paw inflammation. Examination of many neutrophil granule mediators in spinal cord indicated that they are not regulated to the same degree as S100A8 and S100A9. Neutrophil migration also occurs in the vasculature of brain and pituitary gland during peripheral inflammation. Together, these findings suggest an interaction between a subpopulation of leukocytes and the CNS during peripheral tissue inflammation, as implied by an apparent release and possible diffusion of S100A8 and S100A9 through the endothelial blood–brain barrier. Although the present findings do not establish the neurophysiological or behavioral relevance of these observations to nociceptive processing, the data raise the possibility that selective populations of leukocytes may communicate the presence of disease or tissue damage from the periphery to cells in the central nervous system.

Comment on Papers by Chong et al., Nishio et al., and Suri et al. on Diabetes Reversal in NOD Mice

Chong et al., Nishio et al., and Suri et al. (Reports, 24 March 2006, pp. 1774, 1775, and 1778) confirmed that treating nonobese diabetic (NOD) mice with an immune adjuvant and semisyngenic spleen cells can reverse the disease but found that spleen cells did not contribute to the observed recovery of pancreatic islets. We show that islet regeneration predominately originates from endogenous cells but that introduced spleen cells can also contribute to islet recovery.

Susceptibility of dopamine D5 receptor targeted mice to cysteamine

BACKGROUND Recently we demonstrated that gastric mucosa of rats can synthesize, store and release dopamine. Out of five different subtypes, mRNA of D5 (=D1b) dopamine receptor is very abundant in the gastric epithelium. D1 receptor selective dopamine agonists have been shown to protect against experimental gastro-duodenal lesions. AIMS To test the hypothesis that protective effects of dopamine involve D5 receptors, mucosal lesions were induced in D5 receptor deficient (KO) and wild-type (WT) mice using cysteamine. Morphology and gastric acid secretion of D5 KO mice were also studied. METHODS Single doses of 600 mg/kg, 300 mg/kg cysteamine or vehicle were administered subcutaneously to fasted animals. After 24 h, number and severity of gastro-duodenal lesions were analyzed. Basal and histamine-induced maximal gastric acid output were measured by a stomach-sac wash-through method. RESULTS All the KOs in the 600 mg/kg cysteamine group died within 4 h showing symptoms of toxicity while three out of four WTs survived (P<0.05). Mortality after 300 mg/kg cysteamine was significantly higher in KOs versus the WTs: 6/14 versus 2/11, P<0.05. Gastric lesion-index was also significantly higher in KOs (median, middle quartile): four (3-9) versus 0 (0-0), P<0.05. Duodenal lesions did not develop from this single dose of cysteamine in either genotype. Basal and histamine-induced maximal gastric acid output were comparable in the two genotypes. CONCLUSIONS This study demonstrates that loss of D5 receptor causes mucosal vulnerability and increased toxicity of cysteamine in genetically manipulated mice. Thus, D5 receptor subtype is indeed likely to be involved in protective effects of dopamine in the stomach.