Myelostimulatory activity of recombinant human interleukin-2 in mice

In a series of studies designed to extend our understanding of interleukin-2 (IL-2) and to study the effect of biologic response modifiers on bone marrow, we observed that administering recombinant human (rH) IL-2 to normal mice resulted in an increase in the frequency of colony-forming units-culture (CFU-C) in bone marrow. In addition, rH IL-2 was able to accelerate host recovery from cyclophosphamide (CTX)- or radiation-induced bone marrow depression and peripheral blood leukopenia. Not only can rH IL-2 accelerate, in a dose-dependent manner, the return of bone marrow, peripheral blood cellularity, and CFU-C frequency to normal levels following cytoreduction by CTX or irradiation, but it also significantly increases CFU-C frequency to greater than normal levels. Furthermore, rH IL-2 can significantly prolong survival of animals receiving a lethal dose of irradiation or CTX. Thus, multiple mechanisms are responsible for the synergistic therapeutic activity associated with rH IL-2 and CTX. rH IL-2 does not act only as an immunomodulatory agent in the presence or absence of suppressor T cells, but also accelerates host recovery from cytoreductive agents, resulting in decreased leukopenia and perhaps resistances to secondary infection. Thus, rH IL-2 plus chemotherapy may increase therapeutic activity against neoplastic disease, not only by adding immune stimulation to the direct antitumor effect of the drug but also by allowing delivery of higher, more effective doses of chemotherapy.     

Ectopic expression of rhombotin-2 causes selective expansion of CD4-CD8- lymphocytes in the thymus and T-cell tumors in transgenic mice

Although the proto-oncogene rhombotin-2 (RBTN-2) is widely expressed in most tissues, it is not expressed in T cells. We investigated the potential for overexpression of RBTN-2 to cause tumors in T cells and other tissues by constructing transgenic mice that expressed RBTN-2 under control of the metallothionein-1 promoter. Despite overexpression of RBTN-2 in all tissues, transgenic mice developed T-cell tumors only, thus indicating that tumorigenesis caused by RBTN-2 is T-cell-specific. Thymic tumors were found between 37 and 71 weeks and were invariably associated with metastasis to nonlymphoid organs. Thymuses from apparently healthy transgenic mice were also examined. In some mice there was an 10-fold increase in the CD4-CD8- thymocyte subset, yet the total number of thymocytes was the same as that in wild-type mice. Thymic homeostasis was maintained by a compensatory reduction in the CD4+CD8+ subset. The expansion of CD4-CD8- thymocytes was associated with increased expression of RBTN-2 and with increased cell proliferation. No differences were found in the proportion of thymocytes undergoing apoptosis in transgenic mice. Furthermore, RBTN-2- induced expansion of CD4-CD8- cells did not block differentiation of these cells. Thymuses with 30% CD4-CD8- cells were essentially monoclonal, indicating that all thymic immunophenotypes were derived from a single clone. Overall, our data are consistent with the following scenario: (1) RBTN-2 expression in T cells causes selective and polyclonal proliferation of CD4-CD8- thymocytes accompanied by a compensatory decrease in other thymocyte subsets; (2) a clone with growth advantage and differentiation potential is selected and populates the thymus; and (3) this clone eventually breaches homeostasis of the thymus, accompanied or followed by metastasis to other organs.     

Modulation of megakaryocytopoiesis by thrombopoietin: the c-Mpl ligand

We have further characterized the biological activities, mechanism of action, and target cell populations of recombinant human and murine thrombopoietin (rhTPO and rmTPO) in in vitro human and murine model systems. Alone, hTPO or mTPO stimulated the maturation of immature murine megakaryoblasts as measured in a single cell assay. The combination of hTPO or mTPO and interleukin-6 (IL-6) resulted in a further increase in megakaryocyte differentiation in this system. Murine TPO stimulated mouse megakaryocyte progenitor development. Human megakaryocyte progenitor development was potentiated by hTPO alone and further augmented in the presence of the early-acting cytokines (IL-3) or kit ligand/stem cell factor (KL/SCF). To further define the mechanism of action of TPO, neutralization studies were performed with antisera to IL-3, granulocyte-macrophage colony-stimulating factor (GM- CSF), IL-1 beta, and IL-11. No diminution in TPO activity was observed in the presence of these antisera. Moreover, because adhesive interactions are known to modulate hematopoiesis, we studied whether hTPO might alter such interactions between human bone marrow (BM) megakaryocytes and human BM stromal fibroblasts. No changes were observed in either megakaryocyte expression of the surface molecules lymphocyte function-associated antigen-1, very late activation antigen- 4, or intercellular adhesion molecule-1 or the adhesion of megakaryocytes to stromal fibroblasts after treatment with the growth factor. Furthermore, no induction of secretion of the cytokines IL-1 alpha, IL-1 beta, GM-CSF, IL-6, granulocyte-CSF, tumor necrosis factor- alpha, transforming growth factor-beta 1, or transforming growth factor- beta 2 by primary human BM megakaryocytes was noted after treatment of the cells with hTPO. To address whether TPO affects very primitive hematopoietic progenitors, we studied the residual cells from the BMs of mice treated with high doses of 5-fluorouracil. Although no effect of mTPO alone was noted on the viability or replication of such primitive murine progenitor populations, the triple combination of IL-3 + KL/SCF + TPO stimulated growth of megakaryocyte progenitors. These results indicate that TPO is a highly lineage-specific growth factor whose primary biological effects are likely to be direct modulation of the growth and maturation of committed megakaryocyte precursors and immature megakaryoblasts.     

FACTOR VIII RELATED ANTIGEN IN CONNECTIVE TISSUE DISEASE PATIENTS AND RELATIVES

This study assayed serum levels of FVIII Rag as a marker ofendothelial injury in patients not only with frank connectivetissue disease but also in those presenting with Raynaud's phenomenonand in families of those with systemic sclerosis. Elevated levelsof FVIII Rag were found in 62% of patients with systemic sclerosis(SS), 38% with systemic lupus erythematosus (SLE), 67% withmixed connective tissue disease (MCTD) and in 17% with primaryRaynaud's phenomenon. Twenty percent of first degree relativesof patients with SS also demonstrated high levels of FVIII Ragand certain antibodies, namely those reacting with U, RNP andthe centromere. The association between elevated FVIII Rag andantibodies linked to Raynaud's and vasculitis lends supportto antibody involvement in pathogenesis. High levels of FVIIIRag in family members may reflect an increased susceptibilityof endothelium to injury particularly since relatives also havea higher frequency of clinical features such as Raynaud's phenomenon. KEY WORDS: Systemic lupus erythematosus, Raynaud's phenomenon, Scleroderma, Mixed connective tissue disease, Vasculitis     

Phase I trial of interleukin-2 after unmodified HLA-matched sibling bone marrow transplantation for children with acute leukemia

Allogeneic bone marrow transplantation (BMT) for advanced acute leukemia is associated with a high risk of relapse. It is postulated that interleukin-2 (IL-2) administered after BMT might induce or amplify a graft-versus-leukemia effect and thereby reduce the relapse rate. To identify an IL-2 regimen for testing this hypothesis, a phase I trial of IL-2 (Roche) was performed in children in complete remission (CR) without active graft-versus-host disease (GVHD) off immunosuppressive agents after unmodified allogeneic matched-sibling BMT for acute leukemia beyond first remission. Beginning a median of 68 days after BMT, 17 patients received escalating doses of induction IL-2 (0.9, 3.0, or 6.0 x 10(6) IU/m2/d representing levels I, II, and III) for 5 days by continuous intravenous infusion (CIV). After 6 days of rest, they received maintenance IL-2 (0.9 x 10(6) IU/m2/d) for 10 days by CIV infusion. Levels I and II were well-tolerated, but, of 6 patients at level III, 1 developed pulmonary infiltrates, 1 developed hypotension (both resolved), and 1 died of bacterial sepsis and acute respiratory distress syndrome. Grade II acute GVHD developed in 1 patient at level I and 1 at level III. The maximum tolerated dose of induction IL-2 was level II. IL-2 induced lymphocytosis, with an increase in CD56+ and CD8+ cells. Ten patients remain in CR at 5+ to 67+ months. Thus, a regimen of IL-2 has been identified that did not induce a high incidence of acute GVHD when administered to children after unmodified allogeneic BMT. Its clinical activity will be assessed in a phase II trial.     

NEUROPSYCHIATRIC MANIFESTATIONS OF SYSTEMIC LUPUS ERYTHEMATOSUS

Forty-six episodes of major neuropsychiatric manifestations of systemic lupus erythematosus occurring in 35 patients over a ten-year period were reviewed. The frequency of central nervous system SLF was lower than that reported from overseas referral centres. Glucocorticosteroid treatment did not precipitate psychiatric problems in this group. Whilst neuropsychiatric recovery was the rule, such features indicated a subgroup of SLE with high short-term mortality.     

GROUP G STREPTOCOCCAL OSTEOMYELITIS OF THE SPINE

Two elderly men, one with definite and one with probable malignancy,presented with severe back pain. Both had osteomyelitis of thelumbar spine due to Group G streptococci which responded tochemotherapy. KEY WORDS: Streptococcus G, Spine, Osteitis, Elderly men     

THYROTROPHIN RESPONSE TO THYROTROPHIN-RELEASING HORMONE IN OPHTHALMIC GRAVES'' DISEASE: CORRELATION WITH OTHER ASPECTS OF THYROID FUNCTION, THYROID SUPPRESSIBILITY AND ACTIVITY OF EYE SIGNS

Thirty-four patients with ophthalmic Graves' disease were investigated by routine thyroid function tests, by measurement of the serum thyroid-stimulating hormone (TSH) response to thyrotrophin-releasing hormone (TRH) and by measurement of the serum triiodothyronine (T₃) levels. The patients could be divided into four groups according to their response to TRH—normal, impaired, absent and exaggerated. Those with normal responses had routine thyroid function tests and serum T₃ levels which in general did not differ from normal control values. Those with impaired and with absent responses showed routine thyroid function tests which approached the hyperthyroid range together with significant elevation of serum T₃ levels. It is suggested that they might represent examples of ‘subclinical T₃ thyrotoxicosis’. Patients with exaggerated responses had routine thyroid function tests near the lower end of the normal range and could be regarded as suffering from ‘subclinical hypothyroidism’. There was a good correlation between a normal TRH response and normal thyroid suppressibility by T₃, and between impaired and absent responses and impaired thyroid suppressibility. It is evident that the TRH test which is safer, shorter and more convenient, can replace the T₃ suppression test in routine clinical practice. The response to TRH also provided information on the activity of the eye signs, a normal response was associated with improving eye signs whereas impaired, absent or exaggerated responses were associated with static signs or actual deterioration. The TRH test is of value in the diagnosis of unilateral exophthalmos since about three-quarters of the patients in this series showed some abnormality. The wide range of TRH response and of circulating thyroid hormone levels is to be expected since the thyroid is not subject to the normal finely-balanced negative feed-back mechanism via TSH.     

M-mode echocardiography in the detection of surgically resectable left ventricular aneurysm

Thirty- nine patients with severe cardiac failure and a previoustransmural myocardial infarct were studied prospectively toassess the ability of M-mode echocardiography to detect resectableleft ventricular aneurysms. Subsequent angiography showed ananeurysm in 23 patients and diffuse left ventricular damagein 16. Nine (57%) of the patients without aneurysm had eitherleft ventricular end-systolic dimensions in excess of 6 cm combinedwith reduced contraction of the base of the left ventricle (fractionalshortening of the level of the mitral valve <20%), or grossreduction of contraction (fractional shortening <15%) regardlessof left ventricular dimension. These abnormalities did not occurin any patient with an aneurysm. In 21 (91%) of the patientswith a left ventricular aneurysm its presence was correctlypredicted either by fractional shortening in excess of 20% (16patients) or gross delay of mitral valve opening (>80 ms16 patients). In one patient without an aneurysm, fractionalshortening was >20% and thus incorrectly suggested the presenceof aneurysm. Only 39% of aneurysms were directly visualized.Therefore M-mode echocardiography can correctly rule out thediagnosis in over 50% of patients without an aneurysm and stronglysuggest the possibility in over 90% of patients with one.     

Corticobasal and ataxia syndromes widen the spectrum of C9ORF72 hexanucleotide expansion disease

Recently, a hexanucleotide (GGGGCC) repeat expansion in the first intron of C9ORF72 was reported as the cause of chromosome 9p21‐linked frontotemporal dementia‐amyotrophic lateral sclerosis (FTD‐ALS). We here report the prevalence of the expansion in a hospital‐based cohort and associated clinical features indicating a wider clinical spectrum of C9ORF72 disease than previously described. We studied 280 patients previously screened for mutations in genes involved in early onset autosomal dominant inherited dementia disorders. A repeat‐primed polymerase chain reaction amplification assay was used to identify pathogenic GGGGCC expansions. As a potential modifier, confirmed cases were further investigated for abnormal CAG expansions in ATXN2. A pathogenic GGGGCC expansion was identified in a total of 14 probands. Three of these presented with atypical clinical features and were previously diagnosed with clinical olivopontocerebellar degeneration (OPCD), atypical Parkinsonian syndrome (APS) and a corticobasal syndrome (CBS). Further, the pathogenic expansion was identified in six FTD patients, four patients with FTD‐ALS and one ALS patient. All confirmed cases had normal ATXN2 repeat sizes. Our study widens the clinical spectrum of C9ORF72related disease and confirms the hexanucleotide expansion as a prevalent cause of FTD‐ALS disorders. There was no indication of a modifying effect of the ATXN2 gene.