The effect of high altitude on platelet counts, thrombopoietin and erythropoietin levels in young Bolivian airmen visiting the Andes

Recognition of thrombosis as a complication of exposure to high altitude has stimulated interest in rheological changes resulting from hypobaric hypoxia. Previous studies of platelet counts at high altitude have yielded conflicting results and have not been studied in conjunction with potential mediating cytokines. We studied the effects of high-altitude exposure on platelet numbers, thrombopoietin (tpo) and erythropoietin (epo) levels in man. A group of 28 volunteers from the Bolivian Airforce stationed at Santa Cruz (600 m altitude) were studied 48 h and 1 week after their ascent to La Paz (3600 m). In addition 105 volunteers based at Santa Cruz for at least 1 year were compared with 175 age- and sex-matched residents at El Alto (4200 m). Platelet counts were measured immediately after sampling and serum samples assayed for tpo and epo. In the ascending group, mean platelet counts were 251×10⁹, 367×10⁹ and 398×10⁹/l at 600 m and following 48 h and 1 week at 3600 m respectively. Mean tpo levels were 132.5, 76 and 92 pg/ml with epo values of 2.98, 11.6 and 7.9 mIU/ml respectively. In the resident populations mean platelet counts were 271×10⁹/l in the low- and 471×10⁹/l in the high-altitude groups. Mean tpo and epo levels measured 69.3 pg/ml and 4.5 mIU/ml respectively at 600 m and 58.5 pg/ml and 5.1 mIU/ml at 4200 m. In conclusion we have demonstrated a significant and sustained elevation in platelet numbers within 48 h of ascent to high altitude. Our findings do not support a role for tpo as a mediator of the increased platelet count. However, these data do not discount epo as a potential candidate. Received: 28 December 1998 / Revised: 13 May 1999 / Accepted: 26 May 1999     

Microbioreactor Arrays for Full Factorial Screening of Exogenous and Paracrine Factors in Human Embryonic Stem Cell Differentiation

Timed exposure of pluripotent stem cell cultures to exogenous molecules is widely used to drive differentiation towards desired cell lineages. However, screening differentiation conditions in conventional static cultures can become impractical in large parameter spaces, and is intrinsically limited by poor spatiotemporal control of the microenvironment that also makes it impossible to determine whether exogenous factors act directly or through paracrine-dependent mechanisms. We detail here the development of a continuous flow microbioreactor array platform that combines full-factorial multiplexing of input factors with progressive accumulation of paracrine factors through serially-connected culture chambers, and further, the use of this system to explore the combinatorial parameter space of both exogenous and paracrine factors involved in human embryonic stem cell (hESC) differentiation to a MIXL1-GFP⁺ primitive streak-like population. We show that well known inducers of primitive streak (BMP, Activin and Wnt signals) do not simply act directly on hESC to induce MIXL1 expression, but that this requires accumulation of surplus, endogenous factors; and, that conditioned medium or FGF-2 supplementation is able to offset this. Our approach further reveals the presence of a paracrine, negative feedback loop to the MIXL1-GFP⁺ population, which can be overcome with GSK-3β inhibitors (BIO or CHIR99021), implicating secreted Wnt inhibitory signals such as DKKs and sFRPs as candidate effectors. Importantly, modulating paracrine effects identified in microbioreactor arrays by supplementing FGF-2 and CHIR in conventional static culture vessels resulted in improved differentiation outcomes. We therefore demonstrate that this microbioreactor array platform uniquely enables the identification and decoding of complex soluble factor signalling hierarchies, and that this not only challenges prevailing strategies for extrinsic control of hESC differentiation, but also is translatable to conventional culture systems.     

Lymphoblastoid cell-produced immunoglobulins: preparative purification from culture medium by hydroxylapatite chromatography

Hydroxylapatite (HA) is a chromatography matrix capable of separating nucleic acid as well as protein species. HA adsorbs biomolecules as a function of the extent and distribution of their surface charge. HA has been evaluated for its ability to differentially retain immunoglobulin molecules, which are planar relative to the generally globular serum proteins, particularly albumin, contained in tissue culture medium. HA chromatography provides a single-step method to purify and concentrate immunoglobulin proteins secreted by lymphoblastoid cells into culture medium from the vast excess of serum proteins used to supplement the medium. A human lambda light-chain protein was recovered in 5% of the applied volume of medium, and was separated from 95% of the total protein. More than 75% of a hybridoma-produced complete immunoglobulin was recovered essentially free of serum protein contamination. HA appears to provide a valuable alternative chromatographic medium for the purification of immunoglobulin proteins secreted by lymphoblastoid cells.     

Distribution and habitat of Nitellopsis obtusa (Characeae) in the Laurentian Great Lakes

Nitellopsis obtusa, a macroalga (Characeae) native to Europe and Asia, was found in U.S. waters of the St. Clair-Detroit River system in 1983, thus extending the range of this taxon into the Laurentian Great Lakes about 850 km from the St. Lawrence River where it was first discovered in North America in 1978. Its occurrence only in water frequented by commercial shipping vessels suggests that it is distributed via this mechanism. In the St. Clair-Detroit River system, N. obtusa was collected with a Ponar grab at four locations, and with a grapnel at one additional location. It was the ninth most frequently found macrophyte and it was most abundant at Belle Isle in the Detroit River, where the mean dry-weight biomass in Ponar samples was 0 g m^-2 in June, 37 g m^-2 in August, and 32 g m⁻² in September. Maximum biomass of this taxon in one Ponar grab at this location was 289 g m^-2 in September. The alga occurred primarily in water of relatively low current velocity (11.3 cm s⁻¹) and in association with Vallisneria americana, Myriophyllum spicatum, Potamogeton richardsonii, Najas flexilis, and Elodea canadensis. Contribution 654, Great Lakes Fishery Laboratory, Ann Arbor, MI 48105, USA Contribution 654, Great Lakes Fishery Laboratory, Ann Arbor, MI 48105, USA     

Immunological evaluation of repeated administration of erythrocyte-entrapped protein to C3H/HeJ mice.

The immunogenicity of heterologous protein entrapped in autologous erythrocytes was evaluated in a mouse system. Multiple intravenous administrations of an erythrocyte-entrapped bovine beta-glucuronidase preparation resulted in little, if any, immunological response to the entrapped protein or erythrocyte carrier. Erythrocyte entrapment also protected protein from circulating antibody elicited by prior subcutaneous sensitization with unentrapped enzyme in adjuvant. Thus erythrocyte entrapment may protect potentially immunogenic agents from immunological surveillance.     

A comparison of the detection thresholds of odour mixtures and their components

It was the purpose of the present study to investigate whetherthe perception of odour mixtures differs from the perceptionof their components with regard to (i) the reliability of detectionthreshold measurements and (ii) the threshold values themselves.Detection thresholds for a variety of unmixed odorants and 3-,6- or 12-component mixtures were determined for a total of 44subjects, tested repeatedly in five separate sessions usingsniff bottles and an ascending staircase method. Threshold determinationsfor the mixtures were found to be as reliable as for the unmixedsubstances, with intra- and interindividual variability in thresholdvalues actually decreasing with increasing stimulus complexity.The four mixtures for which a systematic comparison of the thresholdvalues of components in the mixed and unmixed state was conductedfailed to provide evidence for the hypo-additive effects ofcompromise or compensation, but rather suggested a positive,sub-threshold interaction between components and possibly evenenhancement. Finally, a significant improvement in individualthreshold scores was seen across test sessions, with experiencedpanelists showing the most stable performance.     

The labelled mitosis curve for a population consisting of fast and slowly cycling cells

In studies of tumour growth, and particularly of tumour treatment with phase-specific chemotherapeutic agents, the fraction labelled mitosis technique is frequently used to estimate the kinetic properties of the cell population making up the tumour. We show here that the FLM technique is in principle very insensitive to the behaviour of slowly cycling cells, even if these constitute a large proportion of the total cell population. Furthermore, since the rate of DNA synthesis is frequently lower in slowly growing cells than in those growing rapidly, there is a higher probability of labelling error associated with the former cells. In view of these theoretical and experimental considerations, it is suggested that considerable caution be used when applying the FLM technique to heterogeneous cell populations such as those of solid tumours.