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Biomedical subjects

M Grossman

Publications and source records attributed to M Grossman.

At least 163 records · Page 9Linked to original sources
PubMed1992

Retroviral-mediated gene transfer in human hepatocytes.

BACKGROUND: The ability to modify human hepatocytes genetically is an essential first step in the development of liver-directed ex vivo gene therapy for inherited metabolic disease. The purpose of these studies was to prove that the genome of human hepatocytes can be altered successfully to express foreign genetic material. METHODS: Human hepatocytes were plated at 2 or 4 x 10(6) cells/10 cm Primaria (Falcon, Oxnard, Calif.) plates. Fresh virus from the amphotropic viral producer cell line BAG, containing the Escherichia coli beta-galactosidase gene lacZ, was placed directly onto hepatocyte cultures and quantitative analysis of cells staining positive for the lacZ gene was undertaken. In a different human liver, a variety of viruses from producer cell lines containing clones of the human low-density lipoprotein (LDL) receptor were plated directly on cultures of human hepatocytes, and gene transfer was demonstrated by increased uptake of fluorescent-labeled LDL. RESULTS: Beta-galactosidase production in hepatocytes was assayed histochemically with the chromogenic substrate X-gal. The highest percentage of cells staining positive for expression of enzyme was seen at 4 x 10(6) cells/plate (43.66% +/- 1.02% vs 27.99% +/- 2.31%). Gene transfer was also documented by the uptake of fluorescent-labeled LDL with a variety of different vectors containing the human LDL receptor. CONCLUSIONS: (1) Human hepatocytes can be cultured in vitro and are susceptible to retroviral infection, (2) functional gene transfer is demonstrated by intracellular function of foreign genes, and (3) the level of expression appears dependent on plating density. We conclude that human hepatocytes are suitable targets for genetic manipulation and may play an important role in human gene therapy trials.

Carbocyanines↗
PubMed1991

Long-term improvement of hypercholesterolemia after ex vivo gene therapy in LDLR-deficient rabbits.

Familial hypercholesterolemia (FH) is an inherited disorder in humans that is caused by a deficiency of low density lipoprotein receptors (LDLRs). An animal model for FH, the Watanabe Heritable Hyperlipidemic rabbit, was used to develop an approach for liver-directed gene therapy based on transplantation of autologous hepatocytes that were genetically corrected ex vivo with recombinant retroviruses. Animals transplanted with LDLR-transduced autologous hepatocytes demonstrated a 30 to 50 percent decrease in total serum cholesterol that persisted for the duration of the experiment (122 days). Recombinant-derived LDLR RNA was harvested from tissues with no diminution for up to 6.5 months after transplantation.

Animals↗
PubMed1991

Receptor-mediated gene delivery in vivo. Partial correction of genetic analbuminemia in Nagase rats.

A plasmid (palb3) was constructed containing the structural gene for human serum albumin driven by mouse albumin enhancer-rat albumin promoter elements. Using an asialoglycoprotein-polycation conjugate consisting of asialoorosomucoid coupled to poly-L-lysine, a soluble DNA complex was formed that was capable of targeting specifically to hepatocytes via asialoglycoprotein receptors present on these cells. Groups of Nagase analbuminemic rats were injected with complexed DNA or controls, followed by two-thirds partial hepatectomy to stimulate hepatocyte replication. Using a cDNA probe for the human albumin structural gene, hybridizable sequences were detected in analbuminemic rats treated with complex as determined by Southern blot analysis. Two weeks post-injection, the targeted DNA was found to exist primarily in plasmid form with an average copy number of 1000/diploid cell. Human albumin mRNA was detected by dot-blot hybridization with a specific oligonucleotide cDNA probe and confirmed by RNase protection assay using a vector-specific probe. Circulating human albumin was detected in the serum of palb3-treated Nagase analbuminemic rats by Western blots using an antibody specific for human serum albumin. A time course demonstrated that circulating human albumin was not detectable 24 h after injection, but became measurable at a level of 0.05 micrograms/ml within 48 h and increased in concentration to a maximum of 34 micrograms/ml by 2 weeks post-injection. This level of expression remained stable through 4 weeks after injection and partial hepatectomy.

Animals↗
PubMed1991

Towards liver-directed gene therapy: retrovirus-mediated gene transfer into human hepatocytes.

Liver-directed gene therapy is being considered in the treatment of inherited metabolic diseases. One approach we are considering is the transplantation of autologous hepatocytes that have been genetically modified with recombinant retroviruses ex vivo. We describe, in this report, techniques for isolating human hepatocytes and efficiently transducing recombinant genes into primary cultures. Hepatocytes were isolated from tissue of four different donors, plated in primary culture, and exposed to recombinant retroviruses expressing either the LacZ reporter gene or the cDNA for rabbit LDL receptor. The efficiency of gene transfer under optimal conditions, as determined by Southern blot analysis, varied from a maximum of one proviral copy per cell to a minimum of 0.1 proviral copy per cell. Cytochemical assays were used to detect expression of the recombinant derived proteins, E. coli beta-galactosidase and rabbit LDL receptor. Hepatocytes transduced with the LDL receptor gene expressed levels of receptor protein that exceeded the normal endogenous levels. The ability to isolate and genetically modify human hepatocytes, as described in this report, is an important step towards the development of liver-directed gene therapies in humans.

Adolescent↗
PubMed1991

Sentence comprehension and praxis deficits in Parkinson's disease.

We evaluated the ability of nondemented patients with idiopathic Parkinson's disease (PD) to interpret various aspects of sentences and to perform learned limb and oral gestures. The patients were significantly compromised in their ability to answer simple questions about sentences such as "The eagle chased the hawk that was fast. Which bird was chased?" A discriminant analysis revealed that up to 73% of PD patients differ from control subjects in their ability to perform this task. Patients with PD were also significantly compromised in their gestural performance, and a discriminant analysis indicated that a praxis deficit may be evident in up to 64% of patients. We conclude that language and gestural processing impairments are frequent in patients with PD.

Aged↗
PubMed1991

Applications of a multiphasic growth function to body composition in pigs.

A multiphasic growth function was used to relate growth of body components to phases of total growth for pigs. Each phase of growth was characterized by asymptotic weight, age at maximum gain, and duration. Age at maximum gain and duration were expressed as a ratio and assumed constant for all phases. One application involved weights of total DM predicted directly with a diphasic function and indirectly with monophasic functions of fat-free DM and fat. Another involved weights of carcass side predicted directly with a diphasic function and indirectly with monophasic functions of offal + muscle + bone and fat + skin. Components were grouped on age at maximum gain. There was good agreement for asymptotic weight between body components and phases, and general agreement for age at maximum gain and for duration, except for carcass weights. A multiphasic growth function may provide a way to examine fat-adjusted weight in living animals because growth of fat appears as a late phase in a multiphasic description of total body growth.

Age Factors↗
PubMed1991

Multitrait animal model with genetic groups.

Genetic groups of unknown parents are extended to the multitrait animal model. Computationally feasible mixed model equations are obtained. A strategy to include genetic groups for missing data is proposed. Canonical and triangular transformations can be applied to the multitrait animal model with groups if the transformations can be applied to the same model without groups. Formulations for REML estimation with groups are derived, and the results are almost as feasible as REML estimation without groups. A numerical example is given to illustrate computations of REML formulations.

Algorithms↗
PubMed1991

Multiphasic analysis of growth curves for progeny of a somatotropin transgenic male mouse.

Diphasic functions were applied to growth curves for body weight and for tail length of mice that were progeny of a transgenic male mated to random-bred NMRI females. A group of 20 female and male mice with high (H) body weight at week 12, assumed to be transgenic, and a group of 20 with normal (N) body weight, assumed to be non-transgenic, were selected for comparison. Body weight and tail length were measured about weekly from 3 to 26 weeks of age. Body weight for H mice at week 26 averaged 1.6 (females) times and 1.9 (males) times that of N littermates. The H mice averaged 1.3 times the gain in weight in first phase for N mice; H mice averaged 2.0 times the gain in second phase for N mice. Tail length for H mice at week 26 averaged 1.1 times that of N littermates. The H mice averaged .9 times the gain in length in first phase for N mice; H mice averaged 1.5 times the gain in second phase for N mice. For H mice, larger tail-length gain in second phase more than compensated for smaller gain in first phase. The transgenic effect may be different for body weight than for tail length. For body weight, the effect was continuous over the entire 26 weeks. For tail length, however, the effect was to delay growth of the tail.

Animals↗
PubMed1991

Multiphasic growth and allometry.

Multiphasic growth assumes increase in body weight, or in other body measures, to be a result of more than one growth phase. Therefore, the concept of allometry can be extended from relation between body measures to relation between phases of growth. For two phases of growth, body weight (W) and tail length (L) can be partitioned into W1 + W2 and L1 + L2. Here, W1 and W2 correspond to phases 1 and 2 of weight and L1 and L2 to phases 1 and 2 of length, where each phase is described by a logistic function. Diphasic functions were applied to growth curves for body weight and for tail length of mice that were progeny of a transgenic male mated to random-bred NMRI females. A group of 20 female and male mice with high body weight at week 12, assumed to be transgenic, and a group of 20 with normal body weight, assumed to be non-transgenic, were selected for comparison. Body weight and tail length were measured about weekly from 3 to 26 weeks of age. Allometric relations between phases for weight (W1 and W2) and tail length (L1 and L2) are presented using predicted values based on estimated parameters of the diphasic growth functions. Differences between ages at maximum gain and ratios of duration of phases were analyzed. Growth in second phase of body weight appeared to be unrelated to growth in first phase of body weight and unrelated to growth in tail length. Growth in each phase of tail length appeared to be close to a simple allometric relation with growth in first phase of body weight. It is now feasible to study multiphasic allometric relations of growth between phases of one body measure and between phases of different body measures by comparing estimates of parameters of the multiphasic growth function.

Animals↗
PubMed1990

Clinical implications of increased plasma levels of CD8 in patients with hairy cell leukemia.

Plasma levels of soluble T-suppressor/cytotoxic antigen (sCD8) were measured at diagnosis or before systemic treatment in 69 patients with hairy cell leukemia (HCL). The 49 nonsplenectomized patients were characterized by high concentrations of sCD8 antigen as compared with 17 controls (P less than .0001). The median sCD8 level in non-splenectomized patients was 1,050 U/mL (range: 160 to 2,400 U/mL) and was significantly higher (P less than .0001) than the median of 275 U/mL (range: 20 to 1,080 U/mL) in splenectomized patients. The relationship of sCD8 to clinical response to subsequent interferon alpha (IFN alpha) treatment was analyzed. Patients who showed subsequent hematologic response with normalization of all blood counts had significantly lower levels of sCD8 concentrations at diagnosis than those who did not (P = .0056). Furthermore, normalization of sCD8 during IFN alpha treatment paralleled the achievement of normal counts in peripheral blood, whereas soluble interleukin-2 receptor (sIL-2R) levels remained high in most patients after 12 to 15 months of treatment. We speculate that activation of suppressor/cytotoxic T cells might play a role in myelosuppression, and its modulation during treatment with IFN alpha correlates with normalization in peripheral blood counts.

Adult↗
PubMed1990

The dynamic relationship between low birthweight and induced abortion in New York City. An aggregate time-series analysis.

We use a vector autoregression to examine the dynamic relationship between the race-specific percentage of pregnancies terminated by induced abortion and the race-specific percentage of low-birthweight births in New York City. With monthly data beginning in 1972, we find that induced abortion explains low birthweight for blacks, but not for whites. There is no evidence of feedback from low birthweight to induced abortion. The findings suggest that unanticipated decreases in the percentage of pregnancies terminated by induced abortion would worsen birth outcomes among blacks in New York City.

Abortion, Induced↗
PubMed1990

Widespread dermatophyte infections that mimic collagen vascular disease.

This article reports the cases of two patients in whom a widespread dermatophyte infection mimicked the cutaneous lesions of their underlying collagen vascular disease. Griseofulvin may be associated with an increased incidence of adverse cutaneous reactions in patients with systemic lupus erythematosus. One patient with systemic lupus erythematosus developed erythema multiforme after taking griseofulvin.

Adult↗
PubMed1990

Expression of human adenosine deaminase in mice reconstituted with retrovirus-transduced hematopoietic stem cells.

Recombinant retroviruses encoding human adenosine deaminase (ADA; adenosine aminohydrolase, EC 3.5.4.4) have been used to infect murine hematopoietic stem cells. In bone marrow transplant recipients reconstituted with the genetically modified cells, human ADA was detected in peripheral blood mononuclear cells of the recipients for at least 6 months after transplantation. In animals analyzed in detail 4 months after transplantation, human ADA and proviral sequences were detected in all hematopoietic lineages; in several cases, human ADA activity exceeded the endogenous activity. These studies demonstrate the feasibility of introducing a functional human ADA gene into hematopoietic stem cells and obtaining expression in multiple hematopoietic lineages long after transplantation. This approach should be helpful in designing effective gene therapies for severe combined immunodeficiency syndromes in humans.

Adenosine Deaminase↗
PubMed1990

Temporary amelioration of hyperlipidemia in low density lipoprotein receptor-deficient rabbits transplanted with genetically modified hepatocytes.

Familial hypercholesterolemia is an inherited disease in humans that is associated with coronary artery disease and is caused by a deficiency of the receptor that mediates the internalization of low density lipoprotein (LDL). We have used an animal model for familial hypercholesterolemia, the Watanabe heritable hyperlipidemic (WHHL) rabbit, to design a therapeutic approach for this disease, which attempts to correct the hepatic defect in LDL receptor expression. Hepatocytes were harvested from WHHL rabbits, plated in primary cultures, and exposed to recombinant retroviruses capable of efficiently transferring a functional human LDL receptor gene. Genetically modified cells were harvested and infused into the portal vein of WHHL recipients, who were analyzed for metabolic consequences of human LDL receptor expression. Each animal exhibited a statistically significant decrease in total serum cholesterol 2-6 days after transplantation, with an eventual return to pretreatment levels. Proviral DNA sequences and virus-directed transcripts were detected in liver tissue 24 hr after transplantation. In situ hybridization demonstrated provirus expression in a small population of hepatocytes distributed in periportal sections of the liver. This study illustrates the potential of somatic gene therapy in ameliorating hyperlipidemia associated with familial hypercholesterolemia.

Animals↗