Biomedical subjects

C A Cain

Publications and source records attributed to C A Cain.

At least 19 recordsLinked to original sources

PubMed2003

Microwave-induced thermal imaging of tissue dielectric properties.

A new imaging method, microwave-induced thermal imaging (MITI), was developed to differentiate tissue based on thermal and dielectric properties. Image contrast depends on temporal strain in tissue, which was determined by one-dimensional speckle tracking using a phase-sensitive, correlation-based technique. The underlying mechanisms were analyzed and experimental results on biologic tissue agreed well with theoretical predictions. Because of its strong contrast between water-bearing and lipid-bearing tissue, the technique may enhance existing intravascular ultrasound (IVUS) imaging systems to identify vulnerable arterial plaque.

Adipose Tissue↗

PubMed2001

Comparison of methods for erythroblast selection: application to selecting fetal erythroblasts from maternal blood.

BACKGROUND: Many methods have been employed to obtain fetal cells from maternal blood for prenatal diagnostics, but there has been little work done that compares the efficacy of different methods. This study presents a comparison of two commonly used methods for selecting erythroblasts with selection directly from whole blood. METHODS: Erythroblasts were isolated from maternal blood by either differential lysis or density separation, followed by selection with an antibody to the transferrin receptor. These methods were compared with antibody selection directly from whole blood. The total yield of erythroblasts was determined for each method. RESULTS: Red cell lysis is not recommended because the lysis step cannot be well controlled. Density separation followed by antibody selection works well. However, a faster and simpler method, antibody selection directly from whole blood using Immunicon Ferrofluid and magnetic separators, works as well and has the potential to yield even more cells. CONCLUSIONS: Considering the need for a simple and quick method for selecting fetal cells from maternal blood, we suggest selection directly from whole blood.

Acetazolamide↗

PubMed2001

A novel method for depositing erythroid cells onto glass slides for fetal cell analysis.

BACKGROUND: We have developed a method for selecting erythroblasts from blood, the first step toward identifying fetal cells in maternal blood for diagnostic purposes. Because the selection method results in a large number of positive cells, we needed to develop new methods to deposit the cells onto slides and to modify in situ hybridization procedures to enable detection of fetal cells. METHODS: We utilized Nunc flaskettes to increase the slide surface area available for cell deposition. The ability of erythroid lineage cells to adhere to several surface modifications was examined. In situ hybridization methods were tested to find the best approach that is compatible with these cell preparations. RESULTS: The best glass slide coating for erythroid cells was found to be an antibody to glycophorin A, a red cell surface antigen. We were able to get excellent in situ hybridization signals in cells on flaskettes by modifying fixation and pretreatment parameters. CONCLUSIONS: The methods described here appear to be the best way of attaching a large number of erythroid lineage cells to slides and of detecting them by in situ hybridization.

Antibodies, Monoclonal↗

PubMed2000

High-density hapten labeling and HRP conjugation of oligonucleotides for use as in situ hybridization probes to detect mRNA targets in cells and tissues.

Oligonucleotides that carry a detectable label can be used to probe for mRNA targets in in situ hybridization experiments. Oligonucleotide probes (OPs) have several advantages over cDNA probes and riboprobes. These include the easy synthesis of large quantities of probe, superior penetration of probe into cells and tissues, and the ability to design gene- or allele-specific probes. One significant disadvantage of OPs is poor sensitivity, in part due to the constraints of adding and subsequently detecting multiple labels per oligonucleotide. In this study, we compared OPs labeled with multiple detectable haptens (such as biotin, digoxigenin, or fluorescein) to those directly conjugated with horseradish peroxidase (HRP). We used branching phosphoramidites to add from two to 64 haptens per OP and show that in cells, 16-32 haptens per OP give the best detection sensitivity for mRNA targets. OPs were also made by directly conjugating the same oligonucleotide sequences to HRP. In general, the HRP-conjugated OPs were more sensitive than the multihapten versions of the same sequence. Both probe designs work well both on cells and on formaldehyde-fixed, paraffin-embedded tissues. We also show that a cocktail of OPs further increases sensitivity and that OPs can be designed to detect specific members of a gene family. This work demonstrates that multihapten-labeled and HRP-conjugated OPs are sensitive and specific and can make superior in situ hybridization probes for both research and diagnostic applications.

Biotin↗

PubMed1999

Identification of differentially expressed mRNAs in human fetal liver across gestation.

Differential gene expression, with its precise start and stop times, is believed to be critical for the programmed development of new cells and tissues. Within the developing fetus, one tissue of particular interest is fetal liver. This organ undergoes rapid changes in the pathway toward liver development in utero since it is also the major site of hematopoiesis, until bone marrow hematopoiesis predominates. Believing that patterns would emerge from the bi-weekly large-scale inspection of expressed genes in the fetal liver, we employed differential display reverse transcription-polymerase chain reaction (DDRT-PCR) as ourprimary inspection tool. Using DDRT-PCR, we isolated cDNAs differentially expressed throughout fetal liver development and in adult liver. We displayed approximately 25 000 cDNAs from 10 and 24 week fetal liver and adult liver. From this initial screen, we determined that approximately 0.1-1% of the mRNA population undergoes expression changes. We extracted, purified and sequenced 25 differentially displayed cDNA bands. Fourteen cDNAs had similarities to known genes, while 11 cDNAs were not similar to any characterized gene. The differentially expressed cDNAs from known genes present in fetal liver include alpha-fetoprotein, stem cell factor, erythroid alpha-spectrin, 2,3-bisphosphoglycerate mutase, insulin-like growth factor-2, porphobilinogen deaminase and Mac30. The differentially expressed cDNAs present in adult liver but not in 10 week fetal liver were nicotinamide deaminase, human fibrinogen-related protein and alpha-acid glycoprotein. The majority of differentially expressed genes found during this effort appear to be turned on during organogenesis, however, some genes were found that are apparently turned off completely.

Adult↗

PubMed1995

Dimethylformamide as an enhancer of cavitation-induced cell lysis in vitro.

Polar solvents, including dimethylformamide (DMF), have been investigated as anticancer drugs, but their potential usefulness is constrained by hepatotoxic side effects. The ability to enhance drug cytotoxicity with ultrasound would be valuable in creating locally intense chemotherapy while minimizing effects peripheral to the treatment site. The effects of continuous wave ultrasound (US) (985 kHz; 0.5-2.5 W/cm2) were evaluated on cultured HL-60 human promyelocytic leukemia cells alone and with a noncytotoxic DMF dose (0.11 M). The cells were insonified in a configuration that created no cell lysis without the introduction of albumin-stabilized microbubbles into the exposure chamber. When microbubbles were introduced, US with bubbles induced cell lysis, and the presence of DMF significantly increased the lysis induced by ultrasound with bubbles. The necessary presence of microbubbles for the DMF-US synergism to occur suggests that a likely mechanism is acoustic cavitation, initiated by the presence of microbubbles as nuclei. Detection of subharmonics confirmed the presence of cavitation, and cell lysis was well correlated with the subharmonic amplitude. The results show that albumin-stabilized microbubbles, similar to those currently used as US contrast agents, may provide a significant source of nuclei and improve prospects for cancer therapy using acoustic cavitation. The evidence presented supports the hypothesis that cell damage is due to a sonochemical rather than to a sonomechanical process.

Cell Membrane↗

PubMed1994

Sonochemical activation of hematoporphyrin: an ESR study.

The production of 2,2,6,6-tetramethyl-4-piperidone-N-oxyl by reaction of 2,2,6,6-tetramethyl-4-piperidone (TMPone) with ultrasonically generated active species in oxygenated solutions of hematoporphyrin (Hp) was studied by electron spin resonance spectroscopy. The nitroxide production rate in air-saturated TMPone solutions in phosphate-buffered saline of pH 9.0 was significantly higher in the presence of Hp than in its absence. The enhancement of nitroxide production by Hp was significantly inhibited in the presence of sodium azide or histidine in the solution. The production rate with Hp was doubled by substitution of deuterium oxide, while the rate without Hp increased only modestly. These results suggest that a substantial amount of active oxygen can be generated by ultrasound in aqueous solutions of Hp. Since the production rate was not reduced by mannitol and no nitroxide was produced in nitrogen-saturated solutions, it appears that hydroxyl radicals do not account for a major portion of the active oxygen species which reacted with TMPone to yield a nitroxide.

Electron Spin Resonance Spectroscopy↗

PubMed1992

Direct computation of ultrasound phased-array driving signals from a specified temperature distribution for hyperthermia.

This paper presents a new method which obtains ultrasound hyperthermia applicator phased-array element driving signals from a desired temperature distribution. The approach combines a technique which computes array element driving signals from focal point locations and intensities with a new technique which calculates focal point locations and power deposition values from temperature requirements. Temperature specifications appear here as upper and lower bounds within the tumor volume, and a focal point placement algorithm chooses focal patterns capable of achieving the temperature range objective. The linear algebraic structure of the method allows rapid calculation of both the phased-array driving signals and an approximate temperature field response. Computer simulations verify the method with a spherical section array (SSA) for a variety of temperature specifications and blood perfusion values. This scheme, which applies to any phased-array geometry, completes an essential step in both treatment planning and feedback for hyperthermia with ultrasound phased-array applicators.

Acoustics↗

PubMed1991

A spherical-section ultrasound phased array applicator for deep localized hyperthermia.

Computer simulation shows that a new ultrasound phased-array with nonplanar geometry has considerable potential as an applicator for deep localized hyperthermia. The array provides precise control over the heating pattern in three dimensions. The array elements form a rectangular lattice on a section of a sphere. Therefore, the array has a natural focus at its geometric center when all its elements are driven in phase. When compared to a planar array with similar dimensions, the spherical-section array provides higher focal intensity gain which is useful for deep penetration and heat localization. Furthermore, the relative grating-lobe level (with respect to the focus) is lower for scanned foci synthesized with this array (compared to a planar array with equal center-to-center spacing and number of elements). This could be the key to the realization of phased-array applicator systems with a realistic number of elements. The spherical-section array is simulated as a spot-scanning applicator and, using the pseudo-inverse pattern synthesis method, to directly synthesize heating patterns overlaying the tumor geometry. A combination of the above two methods can be used to achieve the desired heating pattern in the rapidly varying tumor environment.

Computer Simulation↗

PubMed1990

Time-temperature analysis of cell killing of BHK cells heated at temperatures in the range of 43.5 degrees C to 57.0 degrees C.

Baby hamster kidney (BHK) cells were heated at temperatures in the range of 43.5 degrees C to 57.0 degrees C to determine the time-temperature relationship of cell killing. The cells were grown on 0.025 mm thick pieces of mylar to minimize warm-up times. After heating, the cells were plated for the colony formation assay. The endpoints of 1%, 10%, or 90% isosurvival, or the D0 values of the survival curves were used to construct plots of the logarithm of the reciprocol of the exposure time versus the reciprocol of the absolute temperature. The data for each endpoint resulted in a straight line plot, indicating that the time-temperature relationship for cell killing remained constant from 43.5 degrees C to 57.0 degrees C; namely, a 1.8-fold increase in exposure time was required for a 1 degree C decrease in temperature in order to obtain isosurvival. Heated BHK cells were also examined using electron microscopy. The threshold level of altered morphology was the dissociation of polyribosomal structure and the formation of electron-dense granules within the mitochondria. The time-temperature relationship for the induction of this altered morphology was identical to that for the 90% isosurvival endpoint. Hence, the appearance of altered morphology appears to be related to cell killing.

Animals↗

PubMed1989

Ultrasonic reflection mode imaging of the nonlinear parameter B/A. II: Signal processing.

The nonlinear acoustic interaction between a reflected single-frequency sinusoid and a broadband pump waveform propagating in the opposite direction produces phase changes in the probe proportional to the nonlinear parameter B/A in the spatial region of interaction. The instantaneous phase change along the received probe can be expressed as the convolution of the pump waveform with the spatial distribution of B/A along the propagation path over which the pump and reflected probe interact. In theory, the phase modulated sinusoidal probe can be processed (phase detection and deconvolution) to produce an "A-mode" representation of B/A. If the pump is an intense unipolar impulse and the probe a swept-frequency sinusoid, then the pump interacts with the probe at each point along the propagation path at a unique frequency. Thus the phase modulation that carries information about the spatial distribution of B/A can be extracted from the phase spectrum by a simple Fourier transformation analogous to the space to frequency mapping so basic to magnetic resonance imaging. If the impulsive pump is replaced by another swept-frequency sinusoid, then the phase change in the probe due to B/A at a particular point along the propagation path will be spread out for the duration of the pump along the probe. Passage of the received signal through an appropriate matched filter restores spatial coherence to the phase information in the probe so that it can be processed as if the pump were a broadbanded impulse. This approach suggests a means of approaching the design of effective pump waveforms that can resolve a wide range of spatial frequencies in (B/A)(x).

Algorithms↗

PubMed1987

Ca2+-calmodulin regulates the induction and expression of macrophage cytotoxicity.

Ca2+-activated calmodulin (CAM) is known to regulate cellular responses of diverse cell types to external stimuli. To examine the effects of Ca2+ influx and CAM on macrophage (MP) cytotoxicity, we used 51Cr-labeled cells as targets and in vivo or in vitro activated MPs from C57 BL/6 or BALB/c mice as effectors in a 16-h cytotoxicity assay. MPs activated in vivo by administration of vaccinia virus or in vitro with lymphokine (LK) were cytotoxic for a variety of tumor cell lines, including SV3T3, but not for normal 3T3 cells. Addition of verapamil (0.1 mM), a Ca2+ channel blocker, to MPs activated in vivo by vaccinia virus markedly reduced their cytotoxicity for SV3T3 cells. This correlated with an inhibition of Ca2+ uptake by MPs, as measured by 45Ca influx. Chlorpromazine (20 microM), trifluoperazine (20 microM), and W13 (75 microM), inhibitors of CAM activity, also suppressed MP cytotoxicity for SV3T3 cells when added to the assay, suggesting that Ca2+-activated CAM is an integral component in expression of MP cytotoxicity. To further explore the mechanism of MP cytotoxicity, supernatants from activated MPs treated with various pharmacological agents were examined for cytotoxicity. Vaccinia-activated MPs released a soluble factor(s) that was cytotoxic for SV3T3 cells. Resident MPs cultured under the same condition produced no significant cytotoxic activity. As observed with direct MP cytotoxicity, addition of a Ca2+ channel blocker or CAM inhibitors to cultured activated MPs markedly reduced the cytotoxic activity of the supernatants, suggesting an active secretory process. The requirement for Ca2+ and CAM in the expression of MP cytotoxicity was confirmed in an in vitro MP activation system. Resident cultured MPs activated by an LK preparation were reduced in their tumoricidal capability when verapamil or CAM inhibitors were added to the cytotoxicity assay. This correlated with a reduction in lytic activity of the MP culture supernatants. Further, addition of verapamil or CAM inhibitors to resident MP cultures markedly reduced the induction of tumoricidal MPs by LK, suggesting that Ca2+ and CAM are necessary for both the induction and the expression of MP cytotoxicity. Suppression of cytotoxicity of in vitro activated MP cell line B6MP102 with verapamil and CAM inhibitors confirmed that the MP was the cytotoxic cell being modulated by Ca2+-activated CAM.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1986

Effects of in vivo ultrasound hyperthermia on natural killer cell cytotoxicity in the hamster.

The effects of in vivo ultrasound irradiation of the spleen on immunological functions were assessed with an in vitro natural killer (NK) cell cytotoxic assay. Anesthetized hamsters were exposed to 1 MHz ultrasound at intensity levels currently being used clinically for therapeutic diathermy and hyperthermia (1-5 W/cm2, for 500 sec with constant beam scanning). Hyperthermic levels in the spleen ranged from 38-43 degrees C. Significant depression of natural killer (NK) cell activity was seen 4 h after spleen irradiation as compared to sham irradiated and normal animals. A return towards normal levels was observed in experimental groups at 24 h after exposure. Sham and normal animals were not significantly different in NK activity, indicating no significant stress-related immunosuppressive effects due to handling. Differential leukocyte counts taken for each exposure condition showed significant lymphopenia at 4, 8, and 16 h after exposure, near normal levels at 24 h, and complete recovery by 48 h. The number of circulating mononuclear cells at 4 h showed a dose-related suppression as the exposure intensities were increased.

Animals↗

PubMed1985

Effects of microwave exposure on the hamster immune system. IV. Spleen cell IgM hemolytic plaque formation.

Microwave exposure has been reported to affect various components of the immune system. In this study, we examined the effect of a single whole-body exposure of hamsters to microwave (mw) energy (2.45 GHz; 5-25 mW/cm2; 1 h) on the IgM antibody (Ab) response of spleen cells to sheep red blood cells (SRBC). MW-exposed, sham-exposed, and cage-control hamsters were immunized with SRBC and plaque-forming cells (PFC) in spleens assayed using the direct hemolytic plaque assay. In cage-control hamsters the Ab response was highest between days 4 and 5, returning to baseline by day 9. MW exposure (25 mW/cm2 for 1 h) significantly augmented PFC response only on days 4 and 5 postimmunization, causing approximately a 4.3- and 3.5-fold increase over controls, respectively. Exposure to 15 mW/cm2 caused a lesser, but significant increase in PFC. Exposure to intensities below 15 mW/cm2 for 1 h did not produce any increase in Ab response. Immunization with different concentrations of SRBC following 1 h of 25 mW/cm2 MW exposure revealed a stimulation in PFC at all concentrations ranging from 5 X 10(7) to 5 X 10(8) SRBC. Pretreatment of hamsters with MW radiation prior to immunization showed that the animals retained an increased sensitivity to SRBC for as long as 4 days after MW exposure. In contrast, exposure of hamsters to MW energy on different days after immunization showed an effect of the PFC response only if given between 0 and 1 day after immunization. These results suggest that MW exposure augments the primary IgM response to SRBC by affecting some early event in the immune response process. The various possible explanations for this phenomenon are discussed.

Animals↗

PubMed1985

The effect of preparative protocol on the cytoskeleton ultrastructure observed in extracted whole-mount BHK cells.

Extracted BHK cells (baby hamster kidney) were prepared for electron microscopy by air-drying (with Freon 113), critical-point-drying, and freeze-drying. Variations in the drying procedures had a marked effect on the resultant cytoskeleton ultrastructure. Air drying had to be done in a Freon-saturated atmosphere, residual water had to be removed from the dehydrating solutions and carbon dioxide for critical-point-dried specimens, and freeze-drying had to be done at temperatures lower than -90 degrees C. Failure to exercise these precautions resulted in a cytoskeleton ultrastructure artifact, possibly caused by fusion of the cytoskeleton filaments.

Animals↗

PubMed1985

Binding of calmodulin to the microfilament network correlates with induction of a macrophage tumoricidal response.

Induction of mouse peritoneal macrophage cytotoxicity against SV3T3, a line of virally transformed mouse cells correlated with the distribution of cytoplasmic calmodulin in the macrophages. The organization of the cytoskeleton was examined by fluorescent microscopy and by transmission electron microscopy, using immunogold tagging after Triton-X-100 (TX-100) extraction of the macrophages. Macrophages that had been activated to a tumoricidal state in vivo by vaccinia virus or in vitro by lymphokine stimulation displayed cytoskeletal networks that were more extended and weblike than did resident macrophages. The organization of microfilaments and microtubules in the cytoskeleton was displayed by using either anti-actin or anti-tubulin. Immunogold labeling of tumoricidal macrophage cytoskeletons with anti-calmodulin revealed strong binding to the microfilament network and no binding to microtubules. Anti-calmodulin reacted weakly with the cytoskeletal network of resident macrophages, and this was not demonstrably greater than the reaction with normal sheep serum. However, resident macrophages displayed a high density of calmodulin (CAM) associated with unidentifiable structures in the perinuclear region when reacted with anti-calmodulin. These characteristic distributions of CAM in resident and activated macrophages was confirmed by immunofluorescence. The total and cytoskeletal-associated amounts of calmodulin per unit of protein were determined by radioimmune assay and 125I labeling followed by SDS-PAGE. No statistically significant differences were detected between resident and activated macrophages in either the total cell or cytoskeleton fractions. In summary, our results suggest that induction of tumoricidal activity of mouse peritoneal macrophages correlates with the translocation of calmodulin to the microfilament network of the cytoskeleton.

Animals↗

PubMed1984

Effects of microwave exposure on the hamster immune system. III. Macrophage resistance to vesicular stomatitis virus infection.

Exposure of hamsters to microwave (MW) energy (2.45 GHz, 25 mW/cm2, 1 h) resulted in activation of peritoneal macrophages (PM) to a viricidal state restricting the replication of vesicular stomatitis virus (VSV). The PM from MW-exposed hamsters were viricidal as early as 1 day after exposure and remained active for 5 days. Immunization of hamsters with vaccinia virus induced viricidal PM by 3 to 4 days and they remained active for 7 days. To test the hypothesis that thermogenic MW exposure results in the release of endotoxin across the intestinal epithelium which subsequently activates PM, hamsters were injected with lipopolysaccharide (LPS) and their viricidal activity was studied. Lipopolysaccharide in vitro (0.2 microgram) and in vivo (0.5 microgram) activated macrophages to a viricidal state. When administered in vivo, LPS (0.5 microgram) activated macrophages as early as 1 day and the activity remained for 3 days. While MW exposure of PM in vitro failed to induce viricidal activity, exposure of PM to LPS in vitro induced strong viricidal activity. This suggests that the in vivo response of PM to MW is an indirect one, which is consistent with the hypothesis that MW-induced PM viricidal activity may be mediated via LPS. In preliminary experiments, MW exposure resulted in extended survival time for hamsters challenged with a lethal dose of vesicular stomatitis virus, supporting the concept that MW-activated PM may be a useful therapeutic modality.

Animals↗

PubMed1984

Electromagnetic dosimetry in a Crawford cell: 225 to 500 MHz.

A Crawford cell exposure chamber (main section 30 cm square) was used for electromagnetic dosimetry in the 225 to 500 MHz frequency range. Electric field patterns were measured within the cell. No higher-order modes were observed at any of the frequencies studied below 500 MHz, but significant field perturbations were seen at 2450 MHz. Energy absorption in a prolate spheroidal hamster phantom and in saline-filled spheres of various sizes was measured. The normalized whole-body specific absorption rate (SAR) in watts/kg per mW/cm2 increased from 0.11 (at 225 MHz) to 0.42 (at 450 MHz) for the hamster phantom in E polarization, from 0.06 (at 325 MHz) to 0.20 (at 500 MHz) for K polarization, and from 0.03 (at 325 MHz) to 0.12 (at 500 MHz) for H polarization. Although these values exceed predictions from the long-wavelength far-field approximation for power absorption, the measured SARs were corroborated directly using calorimetry. In addition, calorimetric data obtained using saline-filled spheres suggests that absorbed power in a Crawford cell converges to the far-field plane wave theoretical values only for objects occupying less than 20% of the vertical dimension from the septum to the outer wall.

Electromagnetic Fields↗