Biomedical subjects

D S Miller

Publications and source records attributed to D S Miller.

At least 127 records · Page 7Linked to original sources

PubMed1992

The oocyte nucleus isolated in oil retains in vivo structure and functions.

We describe a technique for isolating the nucleus of the giant amphibian oocyte under paraffin oil. The method precludes the losses of small solutes and proteins that accompany isolation of nuclei into aqueous media. An individual oocyte is blotted, placed under oil, punctured near the animal pole and then squeezed to gently extrude the nucleus into the oil, thereby avoiding exposure to any aqueous environment. Light and electron microscopy of the oil-isolated nucleus demonstrate that its in vivo morphology is preserved. We also describe techniques that facilitate the study of nuclear functions under oil. Oil-isolated oocyte nuclei retain many in vivo functions for several hours, including size-selective envelope permeability, RNA synthesis and the ability to break down in response to cdc2/cyclin meiotic maturation promoting factor.

Animals↗

PubMed1991

Tumor size in endometrial cancer.

Tumor size was determined in 142 patients with clinical Stage I endometrial cancer treated primarily by total abdominal hysterectomy, bilateral salpingo-oophorectomy, and lymph node biopsies between July 1979 and August 1988. Only 4% of patients with tumor size less than or equal to 2 cm had lymph node metastasis; this increased to 15% for tumors more than 2 cm and increased further to 35% when the entire uterine cavity was involved (multivariate P = 0.01). Five-year survival was 98% for patients with tumors less than or equal to 2 cm, 84% with tumors more than 2 cm, and 64% with tumors involving the whole uterine cavity (Mantel-Cox P = 0.005). For endometrial cancer patients with Grade 2 tumors and less than one-half myometrial invasion, the risk of lymph node metastasis is often considered too low to justify adjuvant pelvic radiation therapy. This intermediate-risk group is better defined by including tumor size as a prognostic factor. For this subgroup (Grade 2, less than one-half endometrial invasion) there were no lymph node metastasis associated with tumors less than 2 cm, but 18% had nodal disease when tumors were larger than 2 cm. Tumor size is an important prognostic factor that is particularly helpful in directing adjuvant radiation therapy in patients without staging lymph node biopsies.

Female↗

PubMed1991

Stimulation of protein synthesis by internalized insulin.

Previous studies showed that microinjected insulin stimulates transcription and translation in Stage IV Xenopus oocytes by acting at nuclear and cytoplasmic sites (Miller, D.S., 1988, 1989). The present report is concerned with the question of whether hormone, internalized from an external medium, can act on those sites to alter cell function. Both intracellular accumulation of undegraded 125I-insulin and insulin-stimulated 35S-methionine incorporation into oocyte protein were measured. Anti-insulin antiserum and purified anti-insulin antibody were microinjected into the cytoplasm of insulin-exposed cells to determine if insulin derived from the medium acted through internal sites. In cells exposed for 2 h to 7 or 70 nM external insulin, methionine incorporation was stimulated, but intracellular hormone accumulation was minimal and microinjected antibody was without effect. In cells exposed for 24 h, methionine incorporation again increased, but now accumulation of undegraded, intracellular hormone was substantial (2.6 and 25.3 fmol with 7 and 70 nM, respectively), and microinjected anti-insulin antibody significantly reduced the insulin-stimulated component of incorporation; basal incorporation was not affected. For cells exposed to 70 nM insulin for 24 h, inhibition of the insulin-stimulated component was maximal at 39%. Thus under those conditions, about 40% of insulin's effects were mediated by the internal sites. Together, the data show that inhibition of insulin-stimulated protein synthesis by microinjected antibody was associated with the intracellular accumulation of insulin. They indicate that when oocytes are exposed to external insulin, hormone eventually gains access to intracellular sites of action and through these stimulates translation. Control of translation appears to be shared between the internal sites and the surface receptor.

Animals↗

PubMed1991

Quantitative protein changes in metastatic versus primary epithelial ovarian carcinoma.

Primary and metastatic ovarian carcinomas from six patients were obtained during primary exploratory laparotomy. Tumor cells were synthetically radiolabeled with [35S]methionine. Radiolabeled cellular proteins of the primary and metastatic cells were examined by two-dimensional polyacrylamide gel electrophoresis followed by autoradiography. Computer assisted analysis of the resultant autoradiograms revealed that the amounts of only two proteins, p35 and p36, were consistently and significantly decreased in the metastatic tumor cells. No other consistent differences in protein synthesis between primary and metastatic tumors were detected.

Down-Regulation↗

PubMed1991

Prognostic factors in gestational trophoblastic tumors: a proposed new scoring system based on multivariate analysis.

All 1391 patients treated for gestational trophoblastic tumors (invasive mole and choriocarcinoma) at the John I. Brewer Trophoblastic Disease Center of Northwestern University between 1969 (when use of combination chemotherapy for initial treatment of high-risk disease came into general use) and 1988 were evaluated. Univariate and multivariate analyses were used to determine the relative importance of prognostic factors with respect to survival. The overall cure rate was 93% (363/391): 100% for 223 patients with nonmetastatic disease and 83% for 168 patients with metastatic disease. The only patients who died had a clinicopathologic diagnosis of metastatic choriocarcinoma. In addition to presence of metastasis (83% vs 100%, p less than 0.0001) and diagnosis of choriocarcinoma (67% vs 100%, p less than 0.0001), number of metastases (47% if greater than 8 vs 92% if less than or equal to 8, p less than 0.0001), metastases to sites other than the lung or vagina (52% vs 91%, p = 0.0002), and previous failed chemotherapy (46% vs 84%, p = 0.0014) demonstrated independent significant effects on survival in patients with metastatic disease. A Brewer score, based on our multivariate analysis of survival in patients with metastatic disease, provided predictability of outcome (likelihood ratio chi 2 statistic, chi 2 = 49.8) comparable to that with the World Health Organization score (chi 2 = 45.3), both of which, in turn, were better predictors than either the traditional Hammond clinical classification system (chi 2 = 34.4) or the International Federation of Gynecology and Obstetrics stage (chi 2 = 22.9).

Age Factors↗

PubMed1991

Comparative insights into the mechanisms of renal organic anion and cation secretion.

Comparative models have played a major role in defining the mechanisms that enable vertebrate proximal tubules to transport organic anions and cations from the peritubular interstitium to the urine. The unique advantages of these models and their contributions to our understanding of organic anion and cation transport mechanisms are summarized here. Recent studies of the organic anion transport system suggest that transport is coupled to metabolic energy via indirect coupling to the sodium gradient. Organic anions enter the cell across the basolateral membrane in exchange for alpha-ketoglutarate (alpha-KG), and the alpha-KG is returned to the interior via Na-alpha-KG cotransport. Indirect coupling to Na has been demonstrated in both isolated membranes and intact renal epithelial cells of species ranging from marine crustaceans to mammals. This mechanism was shown to drive not only cellular accumulation but also secretory transepithelial fluxes of organic anions. Luminal exit of secreted organic anions appears to be carrier mediated but is, at present, poorly understood, with mediated potential-driven efflux and anion exchange-driven efflux implicated in some species. As for organic anions, the renal clearance of some organic cations approaches the renal plasma flow. Although there is considerable variation in the handling of specific substrates between species, the basic properties of organic cation transport include carrier-mediated potential-driven uptake at the basolateral membrane, intracellular sequestration that reduces the free concentration of the cation, and luminal exit by organic cation-proton exchange. Reabsorptive transport is also observed for some organic cations, but its mechanisms and driving forces are not well understood.

Animals↗

PubMed1991

Indirect coupling of organic anion secretion to sodium in teleost (Paralichthys lethostigma) renal tubules.

Recent findings in both rat and crab indicate that renal accumulation of p-aminohippurate (PAH) across the basolateral membrane can be coupled indirectly to the Na gradient through PAH-glutarate exchange and Na/glutarate cotransport. However, the role of this mechanism in net transepithelial PAH secretion was not examined. Therefore, proximal tubules from Southern flounder kidney were used to assess both the presence of indirect coupling in the fish and its relationship to net secretion. [14C]glutarate uptake by proximal tubular masses was concentrative, Na dependent, and Li inhibitable. Glutarate efflux from preloaded masses was stimulated by addition of PAH to the medium. Thus flounder tubules exhibited both Na/glutarate uptake and glutarate-PAH exchange. Furthermore, steady-state [3H]PAH accumulation was increased 50% by 10-50 microM glutarate, and this increase was abolished by Li, indicating indirect coupling of PAH entry to Na. To determine whether indirect coupling affected net secretion as well as tissue accumulation, the steady-state accumulation of an anionic dye, fluorescein (FL), was measured in individual renal tubules by use of epifluorescence microscopy and video-image analysis. FL accumulated in tubules to levels that were 20-40 times higher than the medium. In most fish, luminal fluorescence was measurably higher than cellular, and uptake in both compartments was markedly reduced by PAH and Li. Moreover, FL accumulation in cells and lumina was increased by 70-100% when 50 microM glutarate was added to the bathing medium. Thus glutarate not only stimulated uphill FL entry into the cells, but also stimulated active secretion into the tubular lumen.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1991

The role of contraception in the development of postmolar gestational trophoblastic tumor.

From January 1974 to June 1988, 299 evaluable patients were referred to the John I. Brewer Trophoblastic Disease Center of Northwestern University Cancer Center for treatment and/or follow-up of a hydatidiform mole (N = 162) or postmolar gestational trophoblastic tumor (N = 137). The type of contraception and other prognostic factors before and after evacuation were correlated with the development of gestational trophoblastic tumor using both univariate and multivariate analysis. There was no relationship between pre-hydatidiform mole contraception and the development of gestational trophoblastic tumor. Oral contraceptives (OCs) were used by 139 patients (46%), barrier methods by 141 patients (47%), intrauterine devices (IUDs) by two patients (1%), and no contraception by 17 patients (6%). The risk of developing gestational trophoblastic tumor was compared between patients using versus not using: OCs--33 versus 57% (P less than .001), barrier methods--53 versus 40% (P = .30), IUD--100 versus 46% (P = .21), and any contraceptive method--43 versus 88% (P less than .001). The dose of estrogens could be determined in 75 patients taking OCs; 14 of 49 (29%) of the patients taking less than 50 micrograms versus nine of 26 (35%) taking 50 micrograms or more developed gestational trophoblastic tumor (P = .78). Stepwise logistic regression analysis demonstrated that the type of contraceptive used was the most important prognostic factor in gestational trophoblastic tumor development (P less than .0001), followed by the occurrence of theca-lutein cysts (P less than .0001), Asian maternal race (P = .02), lesser time from the last menstrual period (P = .005), and greater maternal age (P = .04).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1991

Dual-parameter flow cytometric analysis coupling the measurements of forward-angle light scatter and DNA content of archival ovarian carcinomas of low malignant potential.

Paraffin-embedded archival specimens from 45 cases of ovarian carcinoma of low malignant potential (OCLMP) were analyzed by flow cytometry (FCM) using propidium iodide (PI) staining. Since single-parameter FCM analysis is often deficient in the resolution of subtle near-diploid DNA-aneuploid populations, forward-angle light scatter (FALS) was measured as a second parameter. DNA aneuploidy was identified in 15 cases (33%). In 7 of those 15 cases, aneuploidy was resolved with single-parameter FCM; in the remaining 8 cases, DNA aneuploidy was resolved only following dual-parameter analysis coupling DNA content and FALS. In all 15 cases, a single near-diploid aneuploid population was observed (mean DNA index = 1.2); there were no tetraploid aneuploid cases. The proliferative activity for all 45 cases studied ranged from 1.0% to 8.9%, with a mean of 3.5%. No difference in mean proliferative activity was observed between the aneuploid and diploid tumors (P greater than .05). To exclude the possibility that PI staining artifacts caused the observed aneuploidy, five of the eight cases shown to be aneuploid by dual-parameter analysis were further studied using an alternate DNA-binding dye, DAPI, yielding similar results. To exclude the possibility that contaminating stromal and/or inflammatory cells caused the observed aneuploidy, samples from a subset of the dual-parameter cases were sorted, revealing the aneuploid populations to be composed primarily of tumor nuclei.(ABSTRACT TRUNCATED AT 250 WORDS)

Aneuploidy↗

PubMed1991

Prognostic factors associated with recurrence in clinical stage I adenocarcinoma of the endometrium.

Two hundred sixty-four consecutive patients with clinical stage I endometrial adenocarcinoma who underwent primary surgical therapy between July 1979 and August 1988 were followed prospectively and evaluated for disease recurrence for 8-112 months (mean 51.5). Thirty-three patients (12.5%) developed recurrence or died of disease. In univariate statistical analysis, prognostic factors significantly associated with disease recurrence were as follows: age (mean 68.6 years with versus 60.3 years without recurrence; P = .0001); histology (adenocarcinoma 8.8%, adenosquamous 35.7%, papillary 25%, clear-cell 57.1%; P less than .0001); tumor grade (grade 1, 7.7%, grade 2, 10.5%, grade 3, 36.1%; P less than .0001); depth of myometrial invasion (none 9.8%, less than one-half 7.4%, one-half or greater 29.6%; P = .0001); lymph node status (negative 8.3%, positive 47.6%; P less than .0001); non-nodal extrauterine disease spread (absent 11.0%, present 50%; P = .0003); peritoneal cytology (negative 9.4%, positive 26.3%; P = .004), and tumor size (2 cm or less 7%, greater than 2 cm 17.3%; P = .05). Cervical extension and uterine size had no significant effect on recurrence. Using multivariate analysis, grade 3 tumor (P = .002), advancing age (P = .004), lymph node metastasis (P = .006), and presence of extrauterine disease spread other than lymph node metastasis (P = .038) were the only variables significantly associated with disease recurrence or death. This study supports the new International Federation of Gynecology and Obstetrics surgical staging system for endometrial cancer.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenocarcinoma↗

PubMed1990

Flow cytometric evaluation of epithelial ovarian cancer.

We investigated the prognostic significance of deoxyribonucleic acid content and proliferative activity of tumor cell populations as measured by flow cytometry of the tumor specimens from 115 women with epithelial ovarian cancer. Deoxyribonucleic acid aneuploidy was found in 87 of 115 (76%) of these cancers with a mean deoxyribonucleic acid index of 1.6 and S-phase fraction of 14.7%. The S-phase fraction of the 28 (24%) diploid tumors was 7.0%. Deoxyribonucleic acid ploidy was significantly correlated with survival. S-phase fraction was significantly correlated with ploidy, residual tumor, histology, grade, ascites, time to recurrence, and survival. Diploidy versus aneuploidy were the best discriminating values for deoxyribonucleic acid index and an S-phase fraction of greater or less than 18% for that parameter. Multivariate analysis revealed stage, S-phase fraction, residual tumor, and grade to be independently associated with time to recurrence, and stage, age, S-phase fraction, and largest metastases were factors associated with survival. Deoxyribonucleic acid ploidy did not significantly improve either model. These results suggest that abnormalities of deoxyribonucleic acid content and the proliferative activity of tumor cell populations are reflective of their biologic activity.

Aneuploidy↗

PubMed1990

Study of the selective cytotoxic properties of cationic, lipophilic mitochondrial-specific compounds in gynecologic malignancies.

Cationic lipophilic compounds have a unique cytotoxic mechanism of action which is dependent on mitochondrial-specific localization of these fluorescent dyes. We have demonstrated in vitro that carcinoma cells, which have a higher negative mitochondrial membrane potential than normal cells, have an increased accumulation and retention of two of these compounds. The compounds tested were rhodamine 123 and dequalinium (DECA). After the development of a reproducible murine intraperitoneal (ip) human ovarian cancer model, which maintained the biologic characteristics of the parent cell line, we undertook in vivo evaluation of DECA. Mice with intraperitoneal tumor inoculations were treated with cisplatin, and/or DECA. When compared to cisplatin, a chemotherapeutic agent known to be effective in the treatment of clinical ovarian cancer, DECA was significantly more efficacious and seemed less toxic in the murine model. Cisplatin and DECA used together were possibly synergistic. Cationic lipophilic compounds may prove to be an exciting new class of antineoplastic agents which exploit intracellular, mitochondrial differences between normal cells and cancer cells.

Animals↗

PubMed1990

Natural killer cell-mediated lysis of T cell lines chronically infected with HIV-1.

The susceptibility of HIV-1-infected CD4+ T cell lines to natural killer (NK) cell-mediated lysis was examined. Non-adherent peripheral blood mononuclear cells (PBMC) of healthy adults lysed HUT cells chronically infected with the IIIB or WMJ1 strains of HIV-1 to a significantly greater extent than uninfected HUT cells. In contrast, Sup-T1 cells chronically infected with these two strains of HIV-1 were not lysed to a greater extent than uninfected Sup-T1 cells. Clone A1.25-infected Sup-T1 (A1.25/Sup-T1), derived from IIIB-infected Sup-T1 cells (IIIB/Sup-T1), were susceptible to non-adherent PBMC-mediated lysis, as were A1.25-infected HUT cells (A1.25/HUT). When non-adherent PBMC were depleted of CD16 (Leu-11b)+ NK cells by treatment with anti-Leu-11b plus C, lysis of HIV-1-infected HUT or Sup-T1 cells was reduced to low levels, indicating that the lysis was mediated by NK cells. Expression of HIV antigens on these target cells did not correlate with their susceptibility to NK cell-mediated lysis. Depletion of interferon-alpha (IFN-alpha) producing HLA-DR+ cells from non-adherent PBMC had no effect on the magnitude of NK cell-mediated lysis of IIIB or WMJ1-infected HUT cells. In contrast, lysis of A1.25/Sup-T1 or A1.25/HUT cells required the presence of HLA-DR+ cells. IFN-alpha production appeared to be required for NK cell-mediated lysis of A1.25/Sup-T1 or A1.25/HUT cells, while lysis of HUT cells infected with the WMJ1 or IIIB strains of HIV-1 was IFN-alpha independent. These results indicate considerable variability in the susceptibility of different HIV-1 infected T cell lines to NK cell-mediated lysis and suggest the existence of alternative mechanisms of activation of NK cells for lysis of HIV-1-infected T cell lines.

Acquired Immunodeficiency Syndrome↗

PubMed1990

Sodium-coupled organic anion transport by Cancer borealis urinary bladder.

Recently, p-aminohippurate (PAH) transport by rat renal basolateral membrane (BLM) vesicles was shown to be coupled indirectly to the Na+ gradient through PAH-glutarate exchange and Na(+)-glutarate cotransport. We have examined the mechanism of PAH transport in Cancer borealis urinary bladder, a simple flat-sheet epithelium that is functionally analogous to vertebrate renal proximal tubule. These experiments indicate that crab bladder is capable of both Na(+)-coupled glutarate uptake and PAH-glutarate exchange and that PAH uptake may be coupled to Na+ in the intact tissue. First, glutarate uptake by intact bladder was lithium sensitive, Na(+) dependent, and inhibited by other dicarboxylates. In flux chambers, the bladder exhibited net glutarate secretion and tissue accumulation occurred primarily from the basolateral side. Furthermore, both BLM and brush-border membrane (BBM) vesicles also exhibited lithium-sensitive Na(+)-coupled glutarate uptake. Second, imposition of an in greater than out glutarate gradient markedly stimulated PAH uptake by bladder BLM vesicles, demonstrating PAH-glutarate exchange. In contrast, exchange was absent in BBM vesicles. Third, in intact bladder tissue, external glutarate increased the steady-state tissue-to-medium ratio for PAH from 14 +/- 1 to 19 +/- 1.5. This increase was both lithium inhibitable and Na+ dependent. Thus not only do bladder BLM show all the elements needed for indirect coupling of PAH transport to the Na+ gradient, but indirect coupling to Na+ can also drive uphill PAH transport in the intact epithelium.

Animals↗

PubMed1989

Stimulation of protein synthesis in stage IV Xenopus oocytes by microinjected insulin.

The effects of intracellular insulin on protein synthesis were examined in intact cells and isolated, undiluted cellular components. [35S]Methionine incorporation into protein was measured in Stage IV oocytes from Xenopus laevis maintained under paraffin oil. Radiolabel and insulin were introduced into the cytoplasm by microinjection. After a short delay (approximately 15 min), injected insulin stimulated the rate of methionine incorporation. Stimulation was dose-dependent, increasing with injected doses in the 7-50-fmol range. Neither proinsulin nor insulin-like growth factor 1 were as effective as insulin in stimulating protein synthesis; microinjected epidermal growth factor and the A and B chains of insulin were without effect. When oocyte surface membranes were removed under oil, the resulting cytoplasm-nucleus samples exhibited methionine incorporation rates that were comparable to those found in intact cells. Microinjection of insulin increased rates of methionine incorporation in cytoplasm-nucleus samples; the effects of external (prior to transfer to oil) and internal (microinjection in oil) insulin exposure were additive. Cytoplasm samples (nuclei and surface membranes removed under oil) also synthesized protein and responded to microinjected insulin. However, insulin responses were reduced relative to cells and to cytoplasm-nucleus samples. 125I-Insulin was degraded rapidly after microinjection into oocytes. Degradation occurred in both the nucleus and cytoplasm. Degradation was delayed by injecting bacitracin into the cells and delaying degradation increased the effectiveness of a low dose of injected insulin. Together, the data show that insulin can act at external, nuclear, and cytoplasmic sites to stimulate protein synthesis in Xenopus oocytes. The signaling pathway activated by internal insulin does not involve plasma membrane-generated second messengers and appears to be separate from that activated by external hormone. Finally, although microinjected insulin is degraded rapidly, it is the intact hormone rather than a degradation product that stimulates protein synthesis.

Animals↗

PubMed1989

Normal caffeine consumption: influence on thermogenesis and daily energy expenditure in lean and postobese human volunteers.

Single-dose oral administration of 100 mg caffeine increased the resting metabolic rate of both lean and postobese human volunteers by 3-4% (p less than 0.02) over 150 min and improved the defective diet-induced thermogenesis observed in the postobese subjects. Measurements of energy expenditure (EE) in a room respirometer indicate that repeated caffeine administration (100 mg) at 2-h intervals over a 12-h day period increased the EE of both subject groups by 8-11% (p less than 0.01) during that period but had no influence on the subsequent 12-h night EE. The net effect was a significant increase (p less than 0.02) in daily EE of 150 kcal in the lean volunteers and 79 kcal in the postobese subjects. Caffeine at commonly consumed doses can have a significant influence on energy balance and may promote thermogenesis in the treatment of obesity.

Adult↗

PubMed1989

Organic anion and cation transport in crab urinary bladder.

Crab urinary bladder, a simple, flat-sheet epithelium, is structurally and functionally analogous to vertebrate renal proximal tubule. Like proximal tubule, crab bladder plays an important role in the excretion of potentially toxic, charged metabolites and xenobiotics. Bladders from Cancer borealis secrete monovalent, organic anions and cations in vivo and in vitro. For organic cations, secretion is a two-step process, with mediated and energetically downhill uptake into cells at the serosal membrane and uphill exit at the luminal membrane. The uptake step may be driven by the electrical potential difference across the serosal membrane, the luminal step by organic cation-proton exchange. Monovalent organic anions are also secreted by a separate two-step process. Recent experiments with intact bladder tissue and isolated membrane vesicles show that (as in mammalian proximal tubule) uphill serosal uptake can be coupled indirectly to the Na+ gradient. Organic anion (p-aminohippurate; PAH) uptake is driven by exchange for certain divalent organic anions, e.g., glutarate and alpha-ketoglutarate. The divalent anion gradient (in greater than out) is in turn maintained by Na+-coupled divalent uptake. The PAH exist step at the luminal membrane is mediated and downhill; it may involve anion exchange.

Animals↗

PubMed1989

Failure to thrive. A continuing problem of definition.

The term "failure to thrive" (FTT) has been used for more than 50 years to describe the infant or young child whose growth falls substantially behind that of his or her peers. Currently, the predominant use of FTT is to describe a child's growth failure resulting from caloric or maternal deprivation or both. Despite widespread use, the term FTT lacks a clarified definition. In this study the authors review 22 current reference texts in general pediatrics, pediatric nutrition, and pediatric gastroenterology and 13 recent journal articles addressing FTT. Only 9 of the 22 texts and 9 of the 13 journal articles describe FTT in quantitative terms, but even these references lack consensus on the anthropometric indices used and their criteria for abnormality. The absence of a standard, scientifically credible definition for FTT, which reflects the dynamic nature of this syndrome, continues to produce an ambiguous body of literature on the subject.

Anthropometry↗