Mobilization of early hematopoietic progenitor cells with BB-10010: a genetically engineered variant of human macrophage inflammatory protein- 1 alpha

BB-10010 is a genetically engineered variant of human macrophage inflammatory protein-1 alpha with improved solution properties. We show here that it mobilizes stem cells into the peripheral blood. We investigated the mobilizing effects of BB-10010 on the numbers of circulating 8-day spleen colony-forming units (CFU-S8), CFU-S12, and progenitors with marrow repopulating ability (MRA). A single subcutaneous dose of BB-10010 caused a twofold increase in circulating numbers of CFU-S8, CFU-S12, and MRA 30 minutes after dosing. We also investigated the effects of granulocyte colony-stimulating factor (G- CSF) and the combination of G-CSF with BB-10010 on progenitor mobilization. Two days of G-CSF treatment increased circulating CFU-S8, CFU-S12, and MRA progenitors by 25.7-, 19.8-, and 27.7-fold. A single administration of BB-10010 after 2 days of G-CSF treatment increased circulating CFU-S8, CFU-S12, and MRA even further to 38-, 33-, and 100- fold. Splenectomy resulted in increased circulating progenitor numbers but did not change the pattern of mobilization. Two days of treatment with G-CSF then increased circulating CFU-S8, CFU-S12, and MRA by 64-, 69-, and 32-fold. A single BB-10010 administration after G-CSF treatment further increased them to 85-, 117-, and 140-fold, respectively, compared with control. We conclude that BB-10010 causes a rapid increase in the number of circulating hematopoietic progenitors and further enhances the numbers induced by pretreatment with G-CSF. BB- 10010 preferentially mobilized the more primitive progenitors with marrow repopulating activity, releasing four times the number achieved with G-CSF alone. Translated into a clinical setting, this improvement in progenitor cell mobilization may enhance the efficiency of harvest and the quality of grafts for peripheral blood stem cell transplantation.     

Reactivity of blood pressure to mental arithmetic stress test, stress-test recovery time, and ambulatory blood pressure in hypertensive and normotensive subjects

BACKGROUND: Laboratory mental stress testing and 24 h ambulatory blood pressure monitoring may analyse reactivity of blood pressure during provoked stress and stressful situations in daily-life, respectively. OBJECTIVE: To evaluate whether the responses to a mental stress test and during the stress-test recovery time were associated with ambulatory blood pressure parameters. METHODS: Fifty-two untreated male subjects (22 normotensives and 30 hypertensives) were subjected both to mental arithmetic stress testing and ambulatory blood pressure monitoring. RESULTS: We found a positive correlation between baseline and peak-test blood pressures during the stress test and 24 h blood pressures. Maximal values of systolic and diastolic blood pressures measured during the 24 h were also correlated to the maximal systolic and diastolic blood pressures reached during the stress test ( P < 0.001). We observed no relationship between reactivity during the stress test and 24 h parameters. On the contrary, changes in diastolic blood pressure during the time of recovery from the stress test (expressed as percentage-change scores) were correlated to the 24 h diastolic blood pressure parameters, the diastolic load being the most closely associated variable. CONCLUSION: The absence of relationships between variations in blood pressure during the provoked stress and ambulatory monitoring parameters indicates that reactivity of blood pressure to an acute stress does not predict the 24 h profile. However, the correlation between the maximal blood pressure measured by ambulatory monitoring and that observed during stress testing indicates that the maximal 24 h values may show the extreme blood pressure response (like the one provoked acutely by a laboratory stress test) of an individual subject. The correlation between the percentage-change score during the recovery time of diastolic blood pressure and the 24 h diastolic load could account forr a lower than normal capacity for recovery of subjects with persistently high blood pressures.     

Bacterial lipase and high-fat diets in canine exocrine pancreatic insufficiency: A new therapy of steatorrhea?

BACKGROUND & AIMS: Nutrients and properties of lipases affect survival of lipolytic activity during aboral gastrointestinal transit. Whether different doses and formulations of bacterial lipase and diets affect steatorrhea was tested in pancreatic-insufficient dogs. METHODS: A dose of 0-600,000 IU of powdered and 135,000 and 300,000 IU of liquid bacterial lipase was given with a standard meal to 5 dogs with ligated pancreatic ducts. In 4 dogs, 0 or 300,000 IU (normal 6-hour postprandial amount) of powder bacterial lipase was also given with five meals containing 850 kcal with different nutrient caloric densities (mixture design). Coefficients of fat absorption during 72- hour fecal balance studies were used to assess treatments. RESULTS: With the standard meal, powder bacterial lipase reduced steatorrhea in a dose-dependent manner (P = 0.03), and 135,000 and 300,000 IU of the liquid form decreased steatorrhea more than powder bacterial lipase (P = 0.017 and 0.057, respectively). Coefficients of fat absorption with 300,000 IU of powder bacterial lipase correlated (r2 = 0.79; P < 0.001) with increasing proportions of fat calories in diets. CONCLUSIONS: Liquid bacterial lipase decreases steatorrhea more than powder, and 300,000 IU of powder bacterial lipase ingested with high-fat meals corrects canine pancreatic steatorrhea. The combination of adequate mixing of small amounts (milligrams) of bacterial lipase and high-fat meals abolishes canine steatorrhea and may abolish human pancreatic steatorrhea. (Gastroenterology 1997 Jun;112(6):2048-55)     

Atrial natriuretic peptide and 24 h blood pressure

BACKGROUND: Atrial natriuretic peptide (ANP) is a hormone involved in the cardiovascular modulation of blood pressure and volume homeostasis. OBJECTIVE: To compare ANP levels in normotensives and hypertensives and to correlate ANP levels with ambulatory blood pressure parameters. METHODS: Plasma samples for ANP determination (using a double-antibody radioimmunoassay Kit) were obtained from 33 consecutive subjects (24 hypertensives, nine normotensives) who had rested supine for 30 min. Afterwards, all of the subjects were subjected to 24 h non-invasive blood pressure monitoring. We found no significant difference between the two groups with regard to ANP levels (95.1+/-29 versus 96.9+/-33 pg/ml, in normotensives and hypertensives, respectively). Also, when hypertensive patients were divided according to their family history of hypertension, ANP levels were similar. There was no correlation between the ANP level and the pre-sampling blood pressure or between the ANP level and the following ambulatory blood pressure monitoring parameters: 24 h, diurnal and nocturnal systolic and diastolic blood pressures, systolic and diastolic loads, nocturnal blood pressure reduction and blood pressure variation coefficients. CONCLUSION: Both the pre-sampling blood pressure and ambulatory monitoring results (sustained blood pressures and pressure variations during the 24 h period) do not seem to influence basal ANP levels in patients with hypertension. These data do not account for a role of this peptide in cardiovascular control, in hypertension.     

Arg-Gly-Asp-dependent occupancy of GPIIb/IIIa by applaggin: evidence for internalization and cycling of a platelet integrin

Using indirect immunofluorescence microscopy we examined the distribution and cycling of GPIIb/IIIa after binding to applaggin, a high-affinity Arg-Gly-Asp (RGD)--containing ligand. Resting, unfixed platelets were incubated with applaggin for 30 minutes at 37 degrees C, and bound applaggin was detected by an affinity-purified rabbit anti- applaggin antibody. Examination of intact cells showed a rim pattern for applaggin, consistent with its binding to the platelet surface. Staining of Triton X-100--permeabilized cells showed an intracellular pool of applaggin. Competition of applaggin binding by either AP-2, an anti-GPIIb/IIIa monoclonal antibody (MoAb) that blocks fibrinogen binding, or the synthetic peptide RGDW eliminated both surface and intracellular staining, indicating that applaggin is binding to GPIIb/IIIa in an RGD-dependent manner. Inhibition of platelet activation by PGE1 and theophylline had no effect on the observed staining patterns, indicating that cellular activation is not required for surface binding and subsequent internalization. To evaluate whether occupancy of functional binding sites on GPIIb/IIIa is required for internalization, we used mAb15, an anti-GPIIIa antibody that neither blocks fibrinogen binding nor induces the expression of ligand-induced binding sites on GPIIb/IIIa. In these studies mAb15 was internalized in a manner analogous to both AP-2 and applaggin, showing that occupancy of the RGD binding site is not required to initiate receptor internalization. To estimate the size of the newly internalized pool of applaggin, 125I-applaggin--binding studies were performed. Displacement of bound 125I-applaggin by excess unlabeled applaggin or EDTA showed that at least 17% of bound applaggin was nondisplaceable when binding was performed under conditions permitting membrane flow and internalization. These data indicate that GPIIb/IIIa is internalized in unstimulated platelets independent of cellular activation or occupancy of the functional binding site(s) of GPIIb/IIIa by RGD-containing ligands. Thus, internalization of GPIIb/IIIa may represent a mechanism by which the surface expression of this adhesion receptor is regulated.     

CD34 expression is regulated reciprocally with adhesion molecules in vascular endothelial cells in vitro

Freshly cultured vascular endothelial cells express the CD34 antigen in a diffuse cell surface pattern with some concentration on microvilli. Expression is downregulated with proliferation in continuous culture and undetectable after nine population doublings but can be maintained by restraining cell proliferation and promoting cell contact. Expression of CD34 at the antigen and mRNA levels on early passage cells is rapidly downregulated by interleukin-1 beta (IL-1 beta), interferon-gamma (INF-gamma), and tumor necrosis factor-alpha (TNF- alpha) under conditions in which these ligands upregulate the adhesion molecules: endothelial leukocyte adhesion molecule 1 (ELAM-1) and intracellular adhesion molecule 1 (ICAM-1). This reciprocal pattern of expression and the topographic distribution of CD34 molecules on the lumenal interdigitated microprocesses of adjacent endothelial cells in vivo suggest that CD34 might have a negative modulating role on adhesion functions of endothelia.     

Autonomic function and baroreflex sensitivity during angiotensin-converting enzyme inhibition or angiotensin II AT-1 receptor blockade in essential hypertensive patients.

OBJECTIVE: The influence of ACE-inhibition and angiotensin II ATI receptor blockade on the autonomic function and baroreflex sensitivity was investigated in hypertension. METHODS AND RESULTS: Heart rate variability was assessed in a resting condition by power spectrum analysis to evaluate the low frequency (LF) power, high frequency (HF) power and LF/HF ratio in 19 hypertensive patients and 23 normotensive controls. Moreover, the coherence between the tachogram and the systogram was evaluated, and the baroreflex gain (alphaLF-index), describing the transfer function of variability in the systolic pressure signal to variability in the RR interval, was obtained.Then a 24-h ambulatory blood pressure monitoring was performed.The 19 hypertensive patients were randomized to either enalapril or losartan treatment, and after 2 months were re-submitted to the RR variability and baroreflex study and to blood pressure monitoring. The subjects then crossed to the other antihypertensive treatment and were re-evaluated after an additional two months. No significant difference was found either in LF power and HF power and LF/HF ratio between normotensive and hypertensive subjects whereas a slight though significant difference was observed in the alphaLF-index. In hypertensive patients, both the treatments with enalapril and losartan reduced blood pressure and had no effect on heart rate. No significant change was observed in autonomic balance or in baroreflex sensitivity during the two antihypertensive treatments. CONCLUSIONS: In hypertensive patients, the angiotensin system or bradykinins do not seem to have any modulatory effect on the sympathetic/parasympathetic control of blood pressure and baroreflex sensitivity, in a resting condition. Since heart rates were unchanged by the two antihypertensive treatments despite a significant reduction of blood pressure, a resetting of baroreflex function was observed during both ACE-inhibition and angiotensin II ATI receptor blockade.     

The final destiny of acantholytic cells in pemphigus is Fas mediated

Background  Pemphigus is an autoimmune disease characterized by the formation of intra-epidermal blisters. Patients develop auto-antibodies against desmoglein 1 and 3 proteins and induce acantholysis.
Objective  This work addresses the issue of whether the Fas pathway mediates acantholysis. Furthermore, the possible suppliers of the Fas pathway were investigated.
Methods  Seventeen biopsies of pemphigus patients were studied by haematoxylin and eosin staining, and apoptosis was defined by TUNEL. The expression of Fas, FasL and caspase 3 was studied by in situ hybridization and immunohistochemistry. Cell infiltrates were studied by immunofluorescence with monoclonal anti-CD3, CD4, CD8, CD19 and CD69.
Results  All of the biopsies showed intra-epidermal blisters, acantholytic cells and inflammatory infiltrates. The blisters expressed Fas, FasL and caspase 3. Cell infiltrates were composed of CD8 and a few CD4⁺CD69⁺ cells. Additionally, CD19⁺ cells were detected. Interestingly, the Fas expression was increased in acantholytic cells and perilesional keratinocytes. Incidentally, these cells exhibited apoptotic features. Interestingly, the CD8 cells expressed FasL.
Conclusion  This paper presents the morphological evidence that apoptosis and acantholysis are linked. Therefore, the Fas pathway is associated with CD8 cells in pemphigus lesions.

Conflicts of interest

None declared.