Nicola M. Kouttab

Long-term bone marrow culture and its clinical potential in chronic wound healing

Bone marrow‐derived cells have long been regarded to play a crucial role in the homeostasis of skin. We have previously described the clinical benefit of directly applying autologous bone marrow aspirate and cultured bone marrow cells to recalcitrant chronic skin wounds. The initial response to treatment appears to be vascular in nature with the formation of new blood vessels. The difficulty in consistently growing adequate numbers of cells for delivery to patients was, however, a limiting factor. Here, in a subsequent protocol, we describe an improved bone marrow culture system yielding a reliable growth of bone marrow cells and leading to a greater clinical response. Cells expressing markers of endothelial progenitors including CD133, CD146, and particularly CD14 are enhanced in these cultures. CD14‐isolated cells produced colonies in endothelial cell assays and sprouting in matrigel assays. Angiogenic cytokines, including angiogenin, epithelial neutrophil‐activating protein‐78, growth‐regulated oncogene, growth‐regulated oncogene‐α, Interleukin‐8, CXC16, and monocyte chemoattractant protein‐1, were found to be elevated in these cultures. Administration of improved culture cells to patients with chronic wounds present for >1 year lead to an enhanced clinical response.

Production of human B and T cell growth factors is enhanced by thymic hormones

The thymic preparations thymosin fraction 5 (TF5) and synthetic thymosin alpha 1 (T alpha 1) were examined for their ability to enhance growth factor production by human peripheral blood mononuclear cells (PBMC). The results showed that both TF5 and T alpha 1 were capable of enhancing the production of a B cell growth factor (BCGF-12kD) and T cell growth factor (TCGF; IL-2). Enhancement by T alpha 1 could be obtained at 100-200-fold lower concentrations than that seen with TF5. In contrast, no enhancement of growth factor production was obtained with control preparations of non-thymic tissue extracts at any concentrations used. It was observed that stimulation of BCGF-12kD and IL-2 was most significantly obtained when the PBMC were activated with lectin. Furthermore, no direct effect of thymic hormones on test B and T cells was observed. These observations provide the first direct evidence that production of B cell growth factors can be enhanced by thymic hormones. In addition, these studies suggest that thymic hormones may regulate B cell responses by acting on mature activated T lymphocytes.

Low expression of Abelson interactor-1 is linked to acquired drug resistance in Bcr-Abl-induced leukemia.

The basis for persistence of leukemic stem cells in the bone marrow microenvironment remains poorly understood. We present evidence that signaling cross-talk between α4 integrin and Abelson interactor-1 (Abi-1) is involved in the acquisition of an anchorage-dependent phenotype and drug resistance in Bcr-Abl-positive leukemia cells. Comparison of Abi-1 (ABI-1) and α4 integrin (ITGA4) gene expression in relapsing Bcr-Abl-positive CD34+progenitor cells demonstrated a reduction in Abi-1 and an increase in α4 integrin mRNA in the absence of Bcr-Abl mutations. This inverse correlation between Abi-1 and α4 integrin expression, as well as linkage to elevated phospho-Akt and phospho-Erk signaling, was confirmed in imatinib mesylate -resistant leukemic cells. These results indicate that the α4-Abi-1 signaling pathway may mediate acquisition of the drug-resistant phenotype of leukemic cells.

Thymic hormone modulation of age-induced changes in the induction of graft-versus-host disease by DBA/2J lymphocytes

Aging induces a number of changes in the immune system, including the involution of the thymus which results in the loss of thymic hormone production and alteration in T cell function. One age-dependent change in immune response is the increasing risk of developing acute or chronic form of graft-versus-host disease (GVHD) following bone marrow transplantation as the age of the recipient increases. A murine model of GVHD that has been extensively studied is one in which injection of C57BL/6 spleen cells into unirradiated B6D2F1 mice results in an acute form of GVHD characterized by cytolytic T lymphocytes (CTL), suppressor cells, runting, and occasionally death. In contrast, injection of DBA/2J spleen cells results in a chronic form of GVHD characterized by a lack of CTL and hyperproduction of immunoglobulin and autoantibodies. This study shows that the GVHD response of DBA/2J spleen cells is dependent on the age of the donor DBA/2J mice. If spleen cells from DBA/2J mice older than 3 months are injected into B6D2F1 recipients, CTL and lack of immunoglobulin production indicative of acute GVHD result. Administration of thymosin fraction 5, a collection of thymic hormones, to DBA/2J mice older than 3 months caused spleen cells from these treated mice to give a GVHD response characteristic of the chronic form of GVHD in B6D2F1 recipients. Thus, thymic hormones were able to modulate the changes in GVHD responses of DBA/2 lymphocytes that occur as the mice age. Preliminary fractionation of TF5 has indicated that there are at least two active thymic peptides present in TF5.

Carnitine and Derivatives in Experimental Infections

Energy in humans and higher animals is generated through s-oxidation of long-chain fatty acids, which are transported across the mitochondrial membrane by carnitine (3-hydroxy-4-methyl-ammoniobutanoate). The bulk of body carnitine is found in cardiac and skeletal tissue, and to a lesser extent in various organs such as the liver.1–3 Thus, generating adequate levels of carnitine is an important factor in maintaining normal metabolic processes. 4,5 Indeed, it has been shown that L-carnitine O-palmitoyltransferase deficiency is accompanied by hypoketoic hypoglycemia and cardiomyopathy,6 an observation which may indicate a role for carnitine in cardiac disease.7,9 Metabolic disorders of fatty acids have also been associated with carnitine deficiency. Syndromes and symptoms include liver dysfuction, disorders of the central nervous system, and skeletal muscle weakness.10 In addition to its classical role in energy metabolism, recent studies have provided evidence for an immunomodulatory role for carnitine, particularly in alleviating pathogenic symptoms induced by infectious agents, as discussed below.