Search PubMed⌕ Search

Biomedical subjects

John M Graham

Publications and source records attributed to John M Graham.

69 records · Page 4Linked to original sources
PubMed2002

OptiPrep density gradient solutions for macromolecules and macromolecular complexes.

Any density gradient for the isolation of mammalian cells should ideally only expose the sedimenting particles to an increasing concentration of the gradient solute. Thus they will experience only an increasing density and viscosity, other parameters such as osmolality, pH, ionic strength and the concentration of important additives (such as EDTA or divalent cations) should remain as close to constant as possible. This Protocol Article describes the strategies for the dilution of OptiPrep in order to prepare such solutions for mammalian cells.

Animals↗
PubMed2002

OptiPrep density gradient solutions for nonmammalian organelles.

Any density gradient for the isolation of nonmammalian organelles should ideally only expose the sedimenting biological particles to an increasing concentration of the gradient solute. Thus they will experience only an increasing density and viscosity, other parameters such as osmolality, pH, ionic strength and the concentration of important additives (such as EDTA and DTT) should remain as close to constant as possible. This Protocol Article describes the strategies for the dilution of OptiPrep in order to prepare such solutions for organelles and membranes from nonmammalian sources such as yeast.

Buffers↗
PubMed2002

OptiPrep density gradient solutions for mammalian organelles.

Any density gradient for the isolation of mammalian organelles should ideally only expose the sedimenting biological particles to an increasing concentration of the gradient solute. Thus they will experience only an increasing density and viscosity, other parameters such as osmolality, pH, ionic strength and the concentration of important additives (such as EDTA and DTT) should remain as close to constant as possible. This Protocol Article describes the strategies for the dilution of OptiPrep in order to prepare such solutions for mammalian organelles and membranes.

Animals↗
PubMed2002

Fractionation of Golgi, endoplasmic reticulum, and plasma membrane from cultured cells in a preformed continuous iodixanol gradient.

A continuous iodixanol gradient within the range 0-30% (w/v) iodixanol can resolve the major membrane compartments of the endoplasmic reticulum, Golgi membranes, and plasma membrane from a postnuclear supernatant prepared from a cultured cell homogenate. The precise density range of the gradient and the centrifugation conditions (100,000-200,000 g for 2-16 h) vary with the type of cell and the requirements of the separation. The strategy is widely used to study the processing of proteins within cells.

Animals↗
PubMed2002

Purification of peroxisomes using a density barrier in a swinging-bucket rotor.

In iodixanol, peroxisomes are the densest organelle in the light mitochondrial fraction and are therefore easily separated from the other components (lysosomes, mitochondria, etc.) in a preformed isosmotic continuous gradient. Because of the large difference in density between peroxisomes and the next densest organelle (mitochondria), a density barrier is effective. The resolution of the peroxisomes is far superior than that in sucrose and, unlike in Percoll, there is no contamination from endoplasmic reticulum.

Animals↗
PubMed2002

Isolation of human polymorphonuclear leukocytes (granulocytes) from a leukocyte-rich fraction.

Human peripheral blood polymorphonuclear leukocytes (PMNs) or granulocytes from a leukocyte-rich plasma (LRP) are banded at an interface between two layers of iodixanol. If the denser layer of iodixanol is omitted the PMNs may alternatively be pelleted. The procedure can be adapted to blood from other species by small changes to the density of the two iodixanol layers. The method works optimally with EDTA- or citrate-anticoagulated blood.

Cell Separation↗
PubMed2002

Purification of intact plant protoplasts by flotation at 1g.

From a standard plant tissue digest adjusted to a density of 1.07 g/ml, protoplasts can be harvested by flotation through a low density barrier (1.03 g/ml). The delicate nature of these bodies is suited to this flotation strategy which can be carried out at 1g.

Cell Wall↗
PubMed2002

Fractionation of hepatic nonparenchymal cells.

The majority of parenchymal cells from mammalian liver cells can be removed by very low speed centrifugation (50 g) but a simple low-density barrier (1.096 g/ml) is required to remove the remaining parenchymal cells from the 50-g supernatant which contains all of the lower density nonparenchymal cells. Continuous gradients of Nycodenz can provide satisfactory resolution of Kupffer, stellate, and endothelial cells on an analytical basis but the separation of different cell types is not sufficient preparatively. Flotation through a low-density iodixanol barrier can, however, provide a satisfactory enrichment of the least dense nonparenchymal cell--the stellate cells.

Cell Separation↗
PubMed2002

Formation of self-generated gradients of iodixanol.

The formation of self-generated gradients of iodixanol from a solution of uniform concentration requires the use of vertical or near-vertical rotors. The density profile that is generated depends upon the sedimentation path length of the rotor, centrifugation time, RCF and temperature. Modulation of the starting concentration changes the density range of the gradient. This Protocol Article illustrates the effect of these parameters on gradient shape in a few selected rotors. Because the gradients are formed by the centrifugal field, they are highly reproducible and easy to execute.

Centrifugation, Density Gradient↗
PubMed2002

Preparation of preformed iodixanol gradients.

The Protocol Article describes the strategies for the formation of preformed discontinuous and continuous density gradients. Most of the techniques can be applied to the use of any gradient solute but some of the detailed recommendations on continuous gradients only apply to iodixanol gradients because of variations in viscosity and rates of diffusion between different solute types. Operational variations that require consideration when dealing with specific types of biological particle are also dealt with at the end of this Protocol Article.

Centrifugation, Density Gradient↗
PubMed2002

A locus for bilateral perisylvian polymicrogyria maps to Xq28.

Polymicrogyria (PMG) is one of a large group of human cortical malformations that collectively account for a significant percentage of patients with epilepsy, congenital neurological deficits, and intellectual disability. PMG is characterized by an excess of small gyri and abnormal cortical lamination. The most common distribution is bilateral, symmetrical, and maximal, in the region surrounding the sylvian fissures, and is known as "bilateral perisylvian polymicrogyria" (BPP). Most cases are sporadic, although several families have been observed with multiple affected members, usually following an X-linked inheritance pattern. Here we report the first genetic locus for BPP mapped by linkage analysis in five families. Linkage places the critical region for BPP at Xq28 (LOD score 3.08 in Xq28, distal to DXS8103 by multipoint analysis). We suggest that this region contains a gene that is necessary for correct neuronal organization and that the identification of this gene will both enhance our understanding of normal cortical development and accelerate the identification of other genes responsible for PMG.

Cerebral Cortex↗
PubMed2002

Infants of diabetic mothers are at increased risk for the oculo-auriculo-vertebral sequence: A case-based and case-control approach.

OBJECTIVES: To determine if infants of diabetic mothers (IDM) are at increased risk for dysplastic ears and the oculoauriculo-vertebral spectrum (OAVS). STUDY DESIGN: Cases of IDM with dysplastic external ears seen at Cedars-Sinai Medical Center were combined with case series in medical literature describing similar patients. Data from a large congenital birth defects registry in Spain were analyzed, and odds ratios (OR) for infants born to either a gestational or preconceptionally diabetic mother to have one of the studied malformations were calculated with 95% confidence intervals. RESULTS: Among the 30 patients in the case series, 50.0% (15) had hemifacial microsomia; 46.7% (14) had hearing loss; 33.3% (10) had facial nerve palsy; 33.3% (10) had vertebral anomalies; 36.7% (11) had cardiovascular defects, of which 45% (5) were conotruncal defects; 26.7% (8) had renal anomalies; 13.3% (4) had limb defects (all radial ray hypoplasia); 10% (3) had DiGeorge sequence; 6.7% (2) had laterality defects; and 6.7% (2) had imperforate anus. Within the cases from the birth defects registry, the odds ratio for OAVS in infants of mothers with gestational diabetes mellitus was 2.28 (95% CI, 1.03-4.82, P =.03), and the OR for ear anomalies in these infants was 1.21 (95% CI, 0.94-1.56, P =.13). When infants of mothers with preconceptionally diagnosed type 1 or 2 diabetes were considered, the OR for OAVS was 1.50 (95% CI, 0.08-9.99, P =.49), and the OR for dysplastic ears was 0.94 (95% CI, 0.48-1.81, P =.85). CONCLUSIONS: Our data indicate that OAVS occurs with a higher incidence in IDM than in the general population. Associated problems include hearing loss, athymia, and cardiac, renal, and limb malformations. Therefore, we recommend that an IDM with features consistent with OAVS undergo a workup including hearing evaluation, skeletal survey, echocardiogram, renal ultrasonogram, and immunodeficiency workup if clinically indicated. Furthermore, noting that most of these defects occur in structures of neural crest origin, we hypothesize that poorly controlled maternal diabetes interferes with cephalic neural crest cell migration.

Adult↗
PubMedDate not supplied

Klinefelter syndrome: expanding the phenotype and identifying new research directions.

PURPOSE: The purpose of this study is to summarize new data on etiology and clinical features of Klinefelter syndrome in order to derive research priorities. METHODS: This study was conducted using critical reviews of selective topics, emphasizing less well-recognized clinical findings. RESULTS AND CONCLUSIONS: The phenotype of the prototypic 47,XXY case is well recognized: seminiferous tubule dysgenesis and androgen deficiency. Less well appreciated is the varied expressivity of 47,XXY Klinefelter syndrome, in particular neurological/cognitive perturbations like language and behavioral problems. Effective therapies are available. Reproductive technologies allow 47,XXY men to sire offspring through intracytoplasmic sperm injection (ICSI); however, genetic counseling is complex and success is low. Behavioral and expressive language difficulties are amenable to treatment by androgen therapy and psychological help. Early treatment may be imperative for optimal outcome.

Biomedical Research↗
PubMedDate not supplied

22q13 deletion syndrome: an update and review for the primary pediatrician.

Recent advances in genetic testing can help to provide a specific diagnosis to children born with syndromes that result in congenital anomalies and developmental delay. One such emerging condition is the 22q13 deletion syndrome. With the introduction of subtelomeric fluorescence-in-situ hybridization (FISH) analysis, the 22q13 deletion has become recognized as a relatively widespread and underdiagnosed cause of mental retardation. Primary-care physicians play an important role in the care of children with 22q13 deletion syndrome, from suspecting the diagnosis in a developmentally delayed child through the medical, developmental, and behavioral aspects of their care. Furthermore, they serve as a valuable source of support and advocacy for the family and a resource for other care providers. The remainder of this article addresses the current state of knowledge regarding 22q13 deletion syndrome and offers the primary-care physician a framework in which to provide care and information.

Child↗