Kenneth H. Falchuk

The molecular basis for the role of zinc in developmental biology

Zinc regulates the gene expression machinery. It affects the structure of chromatin, the template function of its DNA, the activity of numerous transcription factors and of RNA polymerases. Hence, it determines both the types of mRNA transcripts synthesized and the rate of transcription itself. Alterations in one or more of these zinc dependent processes have been proposed to account for the proliferative arrest and teratology induced by zinc deficiency. To examine this proposal, studies of zinc during X. laevis development have been initiated. The kinetics of X. laevis oocyte zinc uptake and storage and of zinc utilization during embryogenesis have been examined first. Vitellogenin carries zinc into the oocyte. Ten % of the total zinc (10 ng/egg) remains within the cytosol while 90% (90 ng/egg) is stored in the yolk platelets associated with lipovitellin. The cytosolic pool is the source of the zinc for all newly formed metalloproteins involved in embryo development. The yolk platelet zinc pool is stored for later use during early metamorphosis. It is now possible to examine zinc transfer to molecules, such as e.g. transcription factors, and the role of the metal in their function in development and organogenesis.

Microparticulate-induced phlebitis: its prevention by in-line filtration

Abstract We carried out a double-blind prospective study of the effect of a filter on the incidence of phlebitis associated with intravenous infusion in 541 patients. A total of 277 patients received infusions through intravenous sets with 0.22-μm IVEX-HP filters, and 264 received infusions without filters. Each infusion was evaluated daily for a maximum of three days. The incidence of phlebitis on Days 1, 2, and 3 of the study was 14.3, 31.1, and 27 per cent for patients receiving infusions without filters and 6.8, 9.7, and 11.3 per cent for those receiving infusions through the filters (P<0.001). Thus, the incidence was reduced by approximately two thirds in the patients who received infusions through the IVEX-HP filters. We conclude that infusion-related phlebitis is a pervasive problem in hospitalized patients, and that it is usually caused by microparticulate components that are present in the infusion fluids and can be removed by in-line filtration. (N Engl J Med 1985;312:78–82.)

Effect of Acute Disease and ACTH on Serum Zinc Proteins

The effect of acute disease and ACTH infusion on serum zinc proteins was studied in serums from 156 healthy and diseased subjects. The mean (+/-2 S.D.) zinc content of 20 normal serums was 96 +/- 20 microng per 100 ml. In 87 serums from acutely ill patients the zinc ranged from 92 to 40 microng per 100 ml. The mean values for nearly all categories of disease studied were lower than normal (P is less than 0.001). Chromatography of normal serum on Sephadex G-100 separates two protein fractions, I and II, containing 37.8+/-8.8 and 76+/-10 microng of zinc per 100 ml, respectively. In serum from diseased patients the zinc in fraction I is unaltered whereas that in fraction II decreases to 29.8+/-7.5 microng per 100 ml (P is less than 0.001). ACTH administration reduces secrum zinc from 10 to 60 microng per 100 ml, the decrements being due to changes in the zinc content of fraction II. Thus, ACTH may have an important role in the reduction of zinc content associated with pathologic states.

A role for biliverdin IXα in dorsal axis development of Xenopus laevis embryos

The determinants of Xenopus laevis embryos that act before their first cell division are mandatory for the formation of mRNas required to establish the dorsal axis. Although their chemical identities are unknown, a number of their properties have long been recognized. One of the determinants is present in the cytoplasm and is sensitive to UV light. Thus, exposing stage 1 embryos to either standard 254-nm or, as shown here, to 366-nm UV light during the 0.3–0.4 time fraction of their first cycle inactivates the cytoplasmic determinant. As a consequence, both types of irradiated embryos fail to express dorsal markers, e.g., goosecoid and chordin, without affecting formation of ventral markers, e.g., Vent-1. The developmental outcome is dorsal axis-deficient morphology. We report here that biliverdin IXα, a normal constituent of cytoplasmic yolk platelets, is photo-transformed by irradiation with either 254- or 366-nm UV light and that the transformation triggers the dorsal axis deficiency. When the 254- or 366-nm UV-irradiated embryos, fated to dorsal axis deficiency, are incubated solely with μM amounts of biliverdin, they recover and form the axis. In contrast, incubation with either in vitro photo-transformed biliverdin or biliverdin IXα dimethyl ester does not induce recovery. The results define an approach to produce dorsal axis-deficient embryos by photo-transforming its biliverdin by irradiation with 366-nm UV light and identify an unsuspected role for biliverdin IXα in X. laevis embryogenesis.

[33] Determination of zinc in biological samples by atomic absorption spectrometry

Publisher Summary At present, the most widely employed methods are based on atomic spectrometry, the interaction of analyte atoms with electromagnetic radiation. These procedures fall into three categories: atomic emission, atomic absorption, and atomic fluorescent spectrometry. The features of the most readily available of these techniques— that is, flame and electrothermic atomic absorption spectrometries, their advantages and their application to the analysis of biological samples will be described in the following sections. In addition, methods for preparing zinc-free buffers and reagents, standards, and specific biological fluids and tissues for analysis, are discussed. The analysis of zinc in most biological samples can be carded out effectively by using either flame or electro thermal atomic absorption spectrometry. The availability of the instruments, their cost, ease of use, and range of sensitivities for zinc determination make them the methods of choice for routine use in biological work on this element. Special situations or problems may require the use of other techniques. The complexities of these other techniques, however, limit their usefulness to specialists skilled in the respective fields.

Spontaneous obliteration of pancreaticoduodenal artery aneurysm after retroperitoneal hemorrhage

We present a rare case of aneurysm of the pancreaticoduodenal artery that bled into the region of the head of the pancreas. Computed tomography (CT) demonstrated the presence and extent of the resultant mass (hematoma) but the angiographic examination allowed a specific diagnosis via visualization of the aneurysm. Unlike previously reported cases, the patient was treated conservatively. Follow-up arteriogram and CT showed spontaneous closure of the pancreaticoduodenal artery and aneurysm and complete resolution of the mass in the pancreatic head. These findings suggest that in some cases a conservative approach to a bleeding pancreaticoduodenal artery aneurysm may be indicated.

Xenopus laevis embryo development: arrest of epidermal cell differentiation by the chelating agent 1,10-phenanthroline.

The embryonic epidermis of stage 35 Xenopus laevis embryos is a highly differentiated structure composed of four cell types arranged in a regular architecture. Each type is distinguished by its distinct morphological characteristics. Some cells are ciliated (type 1); others have their surfaces covered by abundant, secreted vesicles of 0.1 μm diameter (type 2), or multiple linear aggregates of spherical subunits on their apical surfaces (type 3) or large secreted vesicles that emanate from prominent apical holes of 1 μm diameter (type 4). In contrast, the macroscopic appearance of embryos exposed to 10 μM 1,10‐phenanthroline (OP) as well as the ultramicroscopic structure and organization of their epidermal cells are markedly altered. The most predominant cells of the embryonic epidermis are undifferentiated and of heterogeneous size. They lack any characteristic morphology and are arranged irregularly. Ghost cells are also identified. The recognizable differentiated cells are decreased in number and present in a scattered arrangement. These are identified as either type 1 or 2 cells but with ciliae that are shorter and thicker than control or with only a few vesicles larger than 0.1 μm in diameter on their surface. No cells with linear aggregates or prominent apical holes are identified. Except for the altered epidermis, the embryos do not develop any other major organs and exhibit axial abnormalities with an average dorso‐anterior index of three. Thus, the chelating agent OP perturbs metal dependent processes essential for terminal differentiation that may likely account for the resultant abnormalities of embryo organogenesis and morphogenesis. Mol. Reprod. Dev. 55:75–82, 2000.

Subunit Composition of the Zinc Proteins α- and β-Lipovitellin from Chicken

Chicken α- and β-lipovitellin are derived from parent vitellogenin proteins and contain four subunits (125, 80, 40, and 30 kDa) and two subunits (125 and 30 kDa), respectively. Metal analyses demonstrate both are zinc proteins containing 2.1 ± 0.2 mol of zinc/275 kDa per α-lipovitellin and 1.4 ± 0.2 mol of zinc/155 kDa per β-lipovitellin, respectively. The subunits of β-lipovitellin, Lv 1 (MW 125 kDa) and Lv 2 (MW 30 kDa), are separated by gel exclusion chromatography in the presence of zwittergent 3–16. Zinc elutes with Lv 1, suggesting that this subunit binds zinc in the absence of Lv 2. The subunits of α- and β-lipovitellin were separated by SDS-PAGE, digested with trypsin, and mapped by reverse-phase HPLC. The peptide maps of the 125-kDa subunits from α- and β-lipovitellin are essentially identical. Similar results are obtained for the 30-kDa subunits of both lipovitellins. The sequences of five and four peptides of the 125-kDa subunit of α- and β-Lv, respectively, and two peptides of the 30-kDa subunit of α- and β-lipovitellin were determined and match those predicted from the gene for vitellogenin II, Vtg II. Comparison of the amino acid composition of the 125- and 30-kDa subunits of α- and β-lipovitellin support the conclusion that they originate from the same gene. The sequences of peptides from the 80- and 40-kDa subunits of α-lipovitellin have not been found in the NCBI nonredundant data bank. The 27-amino acid N-terminal sequence of the 40-kDa protein is 56% similar to the last third of the Lv 1-coding region of the Vtg II gene, suggesting it may come from an analogous region of the Vtg I gene. We propose a scheme for the precursor—product relationship of Vtg I.

Microparticulate-induced phlebitis: Its prevention by in-line filtration

We carried out a double-blind prospective study of the effect of a filter on the incidence of phlebitis associated with intravenous infusion in 541 patients. A total of 277 patients received infusions through intravenous sets with 0.22-micron IVEX-HP filters, and 264 received infusions without filters. Each infusion was evaluated daily for a maximum of three days. The incidence of phlebitis on Days 1, 2, and 3 of the study was 14.3, 31.1, and 27 per cent for patients receiving infusions without filters and 6.8, 9.7, and 11.3 per cent for those receiving infusions through the filters (P less than 0.001). Thus, the incidence was reduced by approximately two thirds in the patients who received infusions through the IVEX-HP filters. We conclude that infusion-related phlebitis is a pervasive problem in hospitalized patients, and that it is usually caused by microparticulate components that are present in the infusion fluids and can be removed by in-line filtration.