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Biomedical subjects

J T Jones

Publications and source records attributed to J T Jones.

At least 19 recordsLinked to original sources

Ontogeny of odorant receptor gene expression in zebrafish, Danio rerio.

We cloned three putative odorant receptor (OR) genes from the zebrafish to use as in situ hybridization probes to follow the temporal patterns of neurons expressing OR genes through a developmental progression from embryo (12 h postfertilization) to adult. The identification of these genes is supported by sequence homology to previously reported ORs and by the morphology and location of labeled cells in in situ hybridization experiments. Cells expressing OR mRNA were first observed in the olfactory placodes between 31 and 38 h after fertilization (fish reared at 26 degrees C). Initially, only single cells were observed to hybridize the probe; the number of labeled cells increased throughout the remainder of embryogenesis and through postembryonic growth and morphogenesis of the olfactory organ. At all ages, the positively hybridizing cells were scattered throughout the olfactory epithelium but not in the nonsensory epithelium of the olfactory organ.

Amino Acid Sequence

Isolation and characterization of a putative collagen gene from the potato cyst nematode Globodera pallida.

A cDNA clone encoding a full length putative collagen has been isolated in a screen of a mixed stage Globodera pallida expression library. Comparison of the deduced amino acid sequence of this molecule with other collagens suggests it is a cuticular collagen and a member of the col-8 subfamily of collagen genes. Northern blots show the gene is expressed specifically in gravid, adult females of the parasite as compared to second (invasive) stage juveniles and virgin females. Preliminary immunocytochemical studies indicate this collagen is present in areas other than the cuticle; these findings and the potential functional role of this collagen are discussed.

Amino Acid Sequence

Tagging ribozyme reaction sites to follow trans-splicing in mammalian cells.

In mammalian cells, genetic instructions are usually revised by RNA splicing before they are translated to proteins. Here we demonstrate that a trans-splicing group I ribozyme can be employed to intentionally modify the sequence of targeted transcripts in tissue culture cells. By analyzing the ribozyme reaction products, we demonstrate that targeted trans-splicing can proceed in murine fibroblasts with high fidelity, providing direct evidence that ribozymes function as anticipated in a therapeutically relevant setting. Trans-splicing is not very specific however, and the ribozyme reacted with and tagged a variety of cellular transcripts with its 3' exon sequence. RNA tagging provides a unique approach to study RNA catalysis in mammalian cells. Such analysis should facilitate the logical development of safe, therapeutic ribozymes that can repair mutant RNAs associated with a variety of inherited diseases.

3T3 Cells

Inorganic mercury chloride-induced apoptosis in the cultured porcine renal cell line LLC-PK1.

HgCl2 is known to be a renal toxin, but its mechanisms of toxicity are not well understood. The cell line LLC-PK1 was used as a model for renal proximal tubule cells, and the effects of different concentrations of HgCl2 were studied. Apoptosis in response to 35 microM HgCl2 was confirmed by observation of morphological features characteristic of apoptotic cells as well as cleavage of chromosomal DNA into fragments of multiples of 200 base pairs. Ten percent of LLC-PK1 cells in a monolayer underwent apoptosis. These cells detached from the culture flask before apoptosis. Measurement of transepithelial resistance (TER) was used as a functional assay of junctional complex integrity in a novel approach to characterize preapoptotic events in this cell line. Monolayers of LLC-PK1 cells that contained apoptotic cells showed a transient decrease in TER followed by a recovery of TER to the initial levels. The decrease in TER was accompanied by a loss of hemicysts within the monolayer. These data indicate a temporary loss of junctional complexes within the monolayer during apoptosis. One hundred micromolar HgCl2 caused all cells to become necrotic within 3 hr. HgCl2 (10 microM) caused some changes in cell morphology, but no cell death.

Animals

Developmental expression of a turgor-responsive gene that encodes an intrinsic membrane protein.

We previously reported that the pea (Pisum sativum) gene, Trg31, shows increased transcription and elevated mRNA levels in plant tissues which are dehydrated and lose turgor. The protein encoded by Trg31 is homologous to members of the MIP intrinsic membrane protein superfamily. Expression of Trg31 was characterized during pea seedling development and in transgenic tobacco using Trg31 promoter::Gus fusions. In pea, Trg31 mRNA abundance was highest in roots followed by flowers, stems and leaves. In roots, Trg31 mRNA levels were highest in non-elongating regions and low in root tips. In dark-grown seedlings, Trg31 mRNA levels were high in stems and illumination caused mRNA abundance in stems to decrease. Histochemical analysis of transgenic tobacco expressing Trg31 promoter::Gus constructs showed high GUS activity in root to shoot and hypocotyl to cotyledon junctions and cotyledons in germinating seedlings. High activity was also observed in the leaf marginal meristem and trichomes. In more mature seedlings, Trg31 promoter activity was observed in the non-elongating portion of the root and in stems especially in the vascular tissue. A gradient of expression was noted in leaf to stem junction zones with highest expression in the younger tissues. Very high expression was observed in stems of flowers and other floral tissues including the calyx, corolla, style, ovules, pods and pollen. This expression pattern suggests that the Trg31 gene product may facilitate transport from sources, through transmitting tissues to sinks.

Aquaporins

A salt- and dehydration-inducible pea gene, Cyp15a, encodes a cell-wall protein with sequence similarity to cysteine proteases.

The pea (Pisum sativum) gene Cyp15a encodes a protein with sequence similarity to cysteine proteases. Expression of Cyp15a was investigated during pea seedling development and in response to environmental stress. Cyp15a shows increased transcription and elevated mRNA levels in plant tissues that are partially dehydrated or treated with 0.6 M mannitol. Cyp15a mRNA levels also increase in seedlings treated with 0.2-0.25 M NaCl or KCl. During development, Cyp15a mRNA levels increase within 6 to 12 h in cotyledons and axes during germination and continue to increase for at least 96 h. Illumination of dark-grown seedlings increased Cyp15a mRNA abundance in elongating and non-elongating stem tissues. GA and ABA, which modulate the abundance of many seed-localized cysteine proteases, did not significantly modulate Cyp15a mRNA levels in stems. The protein encoded by Cyp15a contains a typical amino-terminal secretory targeting domain. This domain is followed by a pro-sequence containing ca. 110 amino acids that is found in other cysteine proteases. Polyclonal antibodies, directed against CYP15a, recognized both the larger pro-form and the cleaved mature form of CYP15a on western blots. Immunolocalization assays indicated that both forms of the protein are located in cell walls of stem cortical cells.

Cell Wall

Disk submissions.

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Periodicals as Topic

The evaluation of five specialized support surfaces by use of a pressure-sensitive mat.

This study used two full-size pressure-sensitive mats to evaluate five patient support surfaces. The mats, containing 1,536 and 2,340 pressure measurement sites, respectively, made it possible to quantify the entire interface pressure distribution of each support surface. Measurements for groups of 64 and 32 subjects were obtained in supine positions of 0 degree, 30 degrees, at maximal head elevations (50 degrees-60 degrees), and at lateral recumbency. Several statistical descriptors were calculated to characterize the interface pressure distributions. The support surfaces tested included: a powered-air overlay (ACUCAIR Continuous-Air-flow System), a low-airloss bed (FLEXICAIR MC3 Low-Airloss Therapy), a continuous lateral-rotation low-airloss bed (RESTCUE Dynamic Air Therapy), an air-fluidized bed (CLINITRON II Air-Fluidized Therapy), and an integrated bed (prototype Advance 2000 Bed). An analysis of variance revealed that the interface pressure was significantly lower and weight-bearing area was significantly higher on the five surfaces, when compared to the standard hospital mattress.

Beds

Epstein-Barr virus (EBV) and Hodgkin's disease in children: incidence of EBV latent membrane protein in malignant cells.

Previous studies have detected EBV DNA by Southern blotting or in situ hybridization in biopsy material from up to 30 per cent of adult cases of Hodgkin's disease. Here we have used monoclonal antibodies specific for the EBV latent membrane protein LMP1 to examine archival material from children with Hodgkin's disease. Material from 74 cases (54 males and 20 females) was examined and 37 (30 males and 7 females) were classified as LMP1-positive in the malignant cells. LMP1 positivity was present in 4/13 (31 per cent) of lymphocyte predominant, 14/36 (39 per cent) of nodular sclerosis, 17/20 (85 per cent) of mixed cellularity, 1/2 (50 per cent) of lymphocyte depletion, and 1/3 (33 per cent) of unclassified subtypes. The positive cases by clinical stage were I 9/22 (41 per cent), II 9/20 (45 per cent), III 11/24 (46 per cent), and IV 8/8 (100 per cent). LMP1 positivity was present in 2/5 (40 per cent) children aged less than 5 years, 12/27 (44 per cent) aged 5-10 years, and 23/42 (48 per cent) aged between 10 and 15 years. The association between EBV and Hodgkin's disease in children thus appeared to be more frequent in patients with mixed cellularity and advanced disease, but examples of EBV-positive tumours were found in all histological subtypes, stages, and ages. Stepwise discriminant function analysis showed that clinical stage IV and mixed cellularity histology are independently associated with LMP1 positivity. These observations indicate that Hodgkin's disease in children is at least as strongly linked to EBV as is the disease in adults.

Adolescent

Closed-chest cardiac stimulation with a pulsed magnetic field.

Magnetic stimulators, used medically, generate intense rapidly changing magnetic fields, capable of stimulating nerves. Advanced magnetic resonance imaging systems employ stronger and more rapidly changing gradient fields than those used previously. The risk of provoking cardiac arrhythmias by these new devices is of concern. In the paper, the threshold for cardiac stimulation by an externally-applied magnetic field is determined for 11 anaesthetised dogs. Two coplanar coils provide the pulsed magnetic field. An average energy of approximately 12 kJ is required to achieve closed-chest magnetically induced ectopic beats in the 17-26 kg dogs. The mean peak induced electric field for threshold stimulation is 213 V m-1 for a 571 microseconds damped sine wave pulse. Accounting for waveform efficacy and extrapolating to long-duration pulses, a threshold induced electric field strength of approximately 30 V m-1 for the rectangular pulse is predicted. It is now possible to establish the margin of safety for devices that use pulsed magnetic fields and to design therapeutic devices employing magnetic fields to stimulate the heart.

Animals

Inhibition of protein synthesis in irradiated larvae of Schistosoma mansoni.

UV-irradiated and gamma-irradiated schistosomula of Schistosoma mansoni induce high levels of resistance to challenge infection in experimental hosts. It was observed that both types of irradiation severely inhibited protein synthesis by the parasite larvae. Schistosomula were treated with the metabolic inhibitor actinomycin D to simulate this effect of irradiation. The ability of these drug-treated larvae to induce immunity was tested in animal protection experiments. Our results suggest that inhibition of protein synthesis may help to generate the enhanced immunogenicity of irradiated schistosomula. In explanation, we propose that irradiated schistosomula may be such potent immunogens because they express antigens in disrupted, abnormal conformations. Inhibition of protein synthesis may both directly create such modified antigens, and also ensure that they persist and accumulate for presentation to the host immune system.

Animals

Association between surface electromyography of human jaw-closing muscle and quantified food breakdown.

Simultaneous recordings of the activity of the masseter and anterior temporalis and of jaw movement were made on 10 healthy dentate volunteers while they chewed roasted peanuts in their habitual manner. The quantity of food taken was altered by varying (a) its total weight (mouthful) between 8 and 1 g, but with a fixed initial particle size and (b) by varying its initial particle size between median sizes of 9.2 and 2.4 mm, but keeping the mouthful constant. Significant differences were found in the peak and mean (r.m.s.) estimates of muscle activity (particularly in the masseter), and the dimensions of jaw movements. These differences were associated with the variation in both the size of the mouthful and of the initial particles but were much greater with changes in the mouthful. A dimensional argument treating both the weight of food in the mouth and initial particle size as volumes showed that initial particle volume had been varied by approximately 50 times as much as mouthful volume and therefore that the mouthful was a far more critical factor in masticatory physiology than was the particle size of the food. The reasons for this probably lie in an understanding of mechanisms of food comminution.

Adult

Turgor-responsive gene transcription and RNA levels increase rapidly when pea shoots are wilted. Sequence and expression of three inducible genes.

Reduction of turgor in pea shoots caused the accumulation of several poly(A) RNAs. cDNA clones derived from three different poly(A) RNAs which accumulate in wilted pea shoots were isolated, sequenced and expression of the corresponding genes examined. Clone 7a encoded a 289 amino acid protein. The C-terminal 180 amino acids of this protein were homologous to soybean nodulin-26. RNA hybridizing to cDNA 7a was abundant in roots, and induced in shoots by dehydration, heat shock and to a small extent by ABA. Hydropathic plots indicate that the protein encoded by cDNA 7a contains six potential membrane spanning domains similar to proteins which form ion channels. Clone 15a encoded a 363 amino acid protein with high homology to cysteine proteases. RNA hybridizing to cDNA 15a was more abundant in roots than shoots of control plants. Dehydration of pea shoots induced cDNA 15a mRNA levels whereas heat shock or ABA treatment did not. Clone 26g encoded a 508 amino acid protein with 30% residue identity to several aldehyde dehydrogenases. RNA hybridizing to cDNA 26g was induced by dehydration of shoots but not roots and heat shock and ABA did not modulate RNA levels. Levels of the three poly(A) RNAs increased 4-6-fold by 4 h after wilting and this increase was not altered by pretreatment of shoots with cycloheximide. When wilted shoots were rehydrated, RNA hybridizing to cDNA 26g declined to pre-stress levels within 2 h. Run-on transcription experiments using nuclei from pea shoots showed that transcription of the genes which encode the three poly(A) RNAs was induced within 30 min following reduction of shoot turgor. One of the genes showed a further increase in transcription by 4 h after dehydration whereas transcription of the other 2 genes declined. These results indicate that plant cells respond to changes in cell turgor by rapidly increasing transcription of several genes. Furthermore, the expression of the turgor-responsive genes varies with respect to the time course of induction and reversibility of the wilting-induced changes.

Amino Acid Sequence

Sequence and transcriptional analysis of the barley ctDNA region upstream of psbD-psbC encoding trnK(UUU), rps16, trnQ(UUG), psbK, psbI, and trnS(GCU).

A 6.25 kbp barley plastid DNA region located between psbA and psbD-psbC were sequenced and RNAs produced from this DNA were analyzed. TrnK(UUU), rps16 and trnQ(UUG) were located upstream of psbA. These genes were transcribed from the same DNA strand as psbA and multiple RNAs hybridized to them. TrnK and rsp16 contained introns; a 504 amino acid open reading frame (ORF504) was located within the trnK intron. Between trnQ and psbD-psbC was a 2.24 kbp region encoding psbK, psbI and trnS(GCU). PsbK and psbI are encoded on the same DNA strand as psbD-psbC whereas trnS(GCU) is transcribed from the opposite strand. Two large RNAs accumulate in barley etioplasts which contain psbK, psbI, anti-sense trnS(GCU) and psbD-psbC sequences. Other RNAs encode psbK and psbI only, or psbK only. The divergent trnS(GCU) located upstream of psbD-psbC and a second divergent trnS(UGA) located downstream of psbD-psbC were both expressed. Furthermore, RNA complementary to psbK and psbI mRNA was detected, suggesting that transcription from divergent overlapping transcription units may modulate expression from this DNA region.

Amino Acid Sequence

Theoretical basis for controlling minimal tumor temperature during interstitial conductive heat therapy.

This paper describes simulation of steady-state intratumoral temperatures achieved by a simple modality of local heat therapy: interstitial treatment with parallel arrays of warmed, conductive heating elements. During "conductive heating" power is directly deposited only in the interstitial probes. Adjacent tissue is warmed by heat conduction. Simulations of interstitial conductive heating involved solution of the bioheat transfer equation on a digital computer using a finite difference model of the treated tissue. The simulations suggest that when the complete temperature distributions for conductive interstitial hyperthermia are examined in detail, substantial uniformity of the temperature distributions is evident. Except for a thin sleeve of tissue surrounding each heating element, a broad, flat central valley of temperature elevation is achieved, with a well defined minimum temperature, very close to modal and median tissue temperatures. Because probes are inserted directly in tumor tissue, the thin sleeve of overheated tissue would not be expected to cause normal tissue complications. The temperature of the heated probes must be continuously controlled and increased in the face of increased blood flow in order to maintain minimum tumor temperature. However, correction for changes in blood flow is possible by adjusting probe temperature according to a feedback control scheme, in which power dissipation from each probe is the sensed input variable. Conductive interstitial heating with continually controlled probe temperature deserves investigation as a technique for local hyperthermia therapy.

Computer Simulation