Biomedical subjects
Publications and source records attributed to H R Matthews.
We report what we believe to be the first case of Crohn's disease involving the cervical oesophagus with an oesophago-pulmonary fistula to the apex of the lung. The proximal location of the disease prevented radical therapy and the patient eventually died from chronic sepsis.
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The interactions of spermine (4+ charge at physiological pH), N1-acetylspermine(3+), spermidine(3+), N1- and N8-acetylsperimidine(2+), putrescine(2+), hexaamminecobalt(3+), and magnesium(2+) with nucleosome core particles have been examined by using thermal denaturation and circular dichroism. Tetra- and triamines were 2-3 times more effective than diamines at stabilizing core particles against thermal denaturation. Secondary effects were also observed, with acetylpolyamines slightly less effective than unmodified polyamines of equivalent charge. Hexaamminecobalt(3+) was less effective than the triamines, while magnesium had essentially no effect. This is surprising since magnesium is more effective than diamines at stabilizing naked DNA. All the cations tested altered the circular dichroism spectra of the core particles in the DNA region (284 nm). The peak at 284 nm was suppressed by tetra- and trivalent compounds to approximately twice the extent of divalent compounds. Magnesium appears to suppress the peak by a lesser extent than the diamines. This indicates that the DNA twist and/or folding is changed by these cations. A plateau of both thermal denaturation and circular dichroism effects was observed at cation concentrations where 30-40% of the total DNA negative charges could be neutralized by the added cations. We suggest that polyamine and histone acetylation function in concert to lower the stability and change the conformation of the nucleosome core, thus facilitating replication and transcription in vivo.
Oesophageal resection for carcinoma has been performed using a two-part left-sided approach with cervical anastomosis in 75 patients, all of whom have been followed up to death or for a minimum of 5 years. Operative mortality was 12 per cent, but long-term survival was 28 per cent for all squamous tumours and 12 per cent for all adenocarcinomas, irrespective of node status. It is suggested that this approach offers significant advantages.
Putrescine (a diamine) raises the thermal denaturation temperature of mononucleosomes but produces only minor changes in the overall shape of the thermal denaturation curve. This is similar to the effect of sodium ions and is consistent with nonspecific binding to the DNA of the nucleosome. At very low levels of spermidine or spermine the same simple rise in thermal denaturation temperature is seen but at higher levels (above 1 microM for total spermidine concentration) the thermal denaturation curve becomes substantially sharper and the premelt region of the curve diminishes in area. The acetylspermidines display intermediate effects. The change in shape of the thermal denaturation curve was resolved into components (R1 and R2) due to mononucleosomes in their original conformation plus a component (T) induced by the presence of spermidine or spermine. The proportion of component T was substantially reduced with acetylspermidine, compared to equivalent concentrations of spermidine. Hence, we suggest that spermidine acetylation in vivo has the potential to partially destabilise the nucleosome structure, possibly in coordination with histone acetylation.
Myxamoebae of Physarum polycephalum, strain Cld, were grown on agar lawns on live bacteria. Myxamoebae were harvested, fixed and stained with propidium iodide. Flow cytometry showed that, as in the case of Physarum plasmodia, there is no G1 phase during rapid exponential growth. However, an apparent G1 phase was observed at the end of exponential growth when the culture arrested with the G1 DNA content for about a day between growth and differentiation. Most myxamoebae differentiated into cysts, but some formed microplasmodia and others appeared to lose DNA. The cysts possessed the G2 phase DNA content and there was an S phase connecting the G1-arrested state with the encysted state. Encystment was blocked by hydroxyurea (HU) suggesting that DNA synthesis is essential for encystment. The natural temporary synchronization in G1 phase may provide the basis of a method for selecting mutants with a conditional block in G2 or M phases.
Excision of giant emphysematous bullae commonly results in a persistent air leak that requires prolonged intercostal drainage and delays recovery. To minimise this we have used Teflon (polytetrafluoroethylene) strips to buttress the suture line and secure pneumostasis. During 1976-84 eight bullae were excised in seven patients. One patient had bilateral staged thoracotomies. All chest drains were removed within eight days (mean 4.5 days) and no patient developed pulmonary complications. At long term follow up (1-9 years, mean 5.5 years) no complications attributable to the Teflon felt have been identified.
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From 1957 to 1976 oesophageal resection for carcinoma was performed in 1119 patients reported to the West Midlands Cancer Registry. The operations were performed on 581 patients by 127 surgeons who averaged three or less resections per annum (the 'occasional' group). These were compared with 538 patients (the 'frequent' group) whose resections were performed by four surgeons who averaged six or more resections per annum. Operative mortality was 39.4 per cent in the 'occasional' group and 21.6 per cent in the 'frequent' group (P less than 0.001). The age adjusted 5-year survival was 11.1 and 15.2 per cent respectively (P less than 0.05) but when the operative deaths were excluded there was no significant difference. We suggest that oesophageal resection for carcinoma should be performed only where there is an acceptably low operative mortality rate.
Histone H4 from growing cells is partially acetylated at lysines 5, 8, 12, and 16. The turnover rate at each of these sites was investigated by pulse-labeling plasmodia of Physarum polycephalum with [3H]acetate for 55 min in either S phase or G2 phase of the cell cycle. Labeled histone H4 was purified and digested with a protease which cleaves on the carboxyl side of arginine residues. The peptide containing the acetylation sites was purified by high-performance liquid chromatography. Subfractions of the peptide were obtained due to differences in acetyllysine content. Each subfraction was subjected to automated Edman degradation and the radioactivity released after each cycle was determined. Histone H4 was acetylated uniformly in vitro and acetylated peptide 1-23 was used as a control. The results show a very striking preference for turnover on lysine-5 in the "low acetyl" subfraction from cells in S phase; the "high acetyl" subfraction showed turnover at all four sites. The peptides labeled in G2 phase showed turnover mainly at positions -8, -12, and -16. The data imply that the patterns of histone acetyl turnover associated with replication and transcription are nonrandom and distinct. The results have implications for nucleosome structure particularly the possible role of lysine-5 in chromosome maturation and for the design of experiments to test chromatin function in vitro.
The effect of Mg2+, putrescine, diaminopropane, N1-acetylspermidine, N8-acetylspermidine, spermidine, and spermine on the thermal denaturation of calf thymus DNA was investigated. As in a previous study with magnesium [W.F. Dove and N. Davidson, (1962) J. Mol. Biol. 5, 467-478], these ligands were found to raise the thermal denaturation temperature of the DNA and to broaden the thermal denaturation curve dramatically at the point where 10 to 20% of the DNA charge had been neutralized. At higher levels of charge neutralization the curves became sharper again. This behavior was due to differential binding of the ligands to single- and double-stranded DNA. The broadening was used to determine the ratio of the association constants of each ligand to the two forms of DNA using either an independent sites model of binding or an excluded sites model. The results show that the primary mode of binding of the ligands to DNA is electrostatic but that important secondary, nonelectrostatic, effects are also present.
Both cGMP and Ca2+ appear to be involved in the process of phototransduction in vertebrate rods, but their precise roles have been the subject of debate. To investigate the role of Ca2+ we have artificially increased the calcium buffering capacity of the rod by using a patch pipet to incorporate the calcium buffer 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) into the rod cytoplasm. In the presence of buffer the Na+-Ca2+ exchange current became greatly slowed, suggesting that the cytoplasmic calcium concentration (Cai) had indeed been buffered substantially. Although the presence of buffer had negligible effect on the rising phase of the light response, it profoundly altered the later behavior. Responses to brief flashes became prolonged and exhibited an overshoot, apparently because the shut-off process was modified. The normal acceleration of time-to-peak with brighter flashes (an early sign of light adaptation) disappeared. Responses to steady adapting illumination took much longer than normal to settle to a steady level, although the final level represented a similar fractional suppression of current. With superimposed test flashes the presence of such adapting illumination caused a more rapid recovery, whereas the presence of calcium buffer slowed the recovery. The results are consistent with the idea that the rapid drop in Cai, which has recently been shown to accompany the light response, is involved in terminating the light response, and that Cai is thereby involved in setting the operating point and sensitivity of phototransduction. From comparison with other work we infer that Cai appears to act, at least in part, by means of control of cGMP phosphodiesterase activity.
Minitracheotomy is a simple percutaneous technique of tracheal cannulation using a small bore tube. It allows efficient tracheobronchial toilet while preserving glottic function. Examples of its use in the intensive therapy unit are described.
The suction-electrode technique was used to monitor the photocurrent of isolated retinal rods from the tiger salamander, by drawing in the light-sensitive outer segment, or sometimes the inner segment. Calcium buffers or other agents were then introduced into the rod cytoplasm by the 'whole-cell patch-clamp' technique. A patch pipette was sealed against the region of the rod protruding from the suction pipette (usually the inner segment), and the membrane patch was ruptured to obtain a whole-cell recording. Several lines of evidence indicated that the pipette contents diffused into the outer segment, and showed that the cell could be adequately voltage clamped. With only trace quantities of chelator in the patch pipette (to bind stray calcium), a gradual decline of the dark current and slowing of responses was usually observed over a period of 10-20 min after rupture of the patch. When the patch pipette contained no added calcium and 10 mM of the calcium chelator BAPTA (1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid) (free Ca2+ ca. 10(-9) M) rupture of the patch led, over a period of a few minutes, to an increase in mean dark current, an increased duration of responses, a substantial increase in flash sensitivity, and a pronounced overshoot in the recovery phase, but with virtually no change in the rising phase of the response to bright flashes. Similar results were obtained when EGTA was used in place of BAPTA, and also in the few cases when successful rupture of the outer segment membrane was obtained. With the free calcium concentration in the patch pipette buffered to the higher level of 1 microM (with 10 mM-Ca2+/11 mM-BAPTA) the results were qualitatively similar to those obtained with BAPTA alone, except that the mean dark current did not increase. This is consistent with a resting free calcium concentration in darkness in the region of 1 microM. In the presence of bright steady illumination with BAPTA in the cell the suppression of outer segment current could be maintained for at least 15 min. Upon extinction of the light a very large current transient developed (similar to the overshoot with flashes) which was light suppressible. With backgrounds of moderate intensity the incorporation of buffer led to a gradual reduction of the residual current.(ABSTRACT TRUNCATED AT 400 WORDS)
Seventeen patients with respiratory failure from a variety of causes have been treated by high-frequency jet ventilation, delivered by a specially designed Y connector and a standard minitracheostomy tube, for periods ranging from 12 hours to 14 days. This percutaneous system provides access for full ventilatory support and tracheobronchial suction, but without the need for a cuffed endotracheal tube or sedation. Patients can eat, drink, and talk during ventilation. Ten of the patients were male and ages ranged from 31 to 77 years. Ventilation was satisfactory in 16 patients and unsatisfactory in one, who had also been difficult to manage on conventional ventilation. Thirteen patients resumed full spontaneous respiration and four died of their underlying disease while still being supported with a ventilator. These results suggest that the combination of minitracheostomy and high-frequency jet ventilation may offer significant advantages in the management of some patients with respiratory failure.
A high salt nuclear extract from the true slime mold Physarum polycephalum was used as a source of kinase activity for the incubation of calf thymus histones with [gamma-32P]ATP. A major proportion of the 32P incorporated into histones was acid-labile and alkali-stable. The nature of the alkali-stable phosphorylated component was analyzed by subjecting the phosphorylated protein to total alkaline hydrolysis and separating the resultant phosphoamino acids by anion exchange chromatography. The 32P-labeled material co-chromatographed with phosphohistidine standards and did not co-chromatograph with phosphoserine, phosphothreonine, or phosphotyrosine standards. In similar experiments using reversed phase high-performance liquid chromatography to separate the phosphoamino acids, the 32P-labeled phosphoamino acid behaved like the 1-isomer of phosphohistidine, in not being retained by the column, and unlike 3-phosphohistidine, phosphoserine, phosphothreonine, phosphotyrosine, and phosphoarginine, which were all retained on the column. Histone H4 was a good substrate for the histidine kinase activity and the location of the phosphorylated histidine residue was probed by peptide mapping using chymotrypsin or V8 protease. Both maps were consistent with labeling of histidine 75 and inconsistent with labeling of histidine 18. The data show that Physarum nuclei contain a major kinase activity which produces phosphohistidine. The methods we have developed for studying this kinase activity provide the basis for a complete characterization of the structure and function of the Physarum enzyme and can be applied to the study of similar kinase activities in other systems.
Amino acids were derivatized with orthophthalaldehyde and separated by high-performance liquid chromatography on a polymer-based reverse-phase column (Hamilton PRP-1) at pH 7.2 using isocratic elution with 14.3 mM sodium phosphate, 1.1% tetrahydrofuran, 6.6% acetonitrile. Phosphorylated amino acids were eluted with baseline resolution in the following order: 1-phosphohistidine, phosphoserine, 3-phosphohistidine, phosphotyrosine, phosphothreonine, and phosphoarginine. Each of the phosphoamino acids was separated from its parent amino acid but aspartate and glutamate eluted in the same region as the phosphoamino acids. The sensitivity is in the picomole range and the separation time, injection to injection, is 15 min. The linearity for phosphothreonine extends at least from 30 pmol to 30 nmol. Quantitation by radioactivity is good for each of the phosphoamino acids except in the case of [1-32P]phosphohistidine, which coelutes with inorganic phosphate.
A computer program is described that emulates a curve analyzer and fits a sum of up to 30 Gaussian components plus a straight background to any set of equi-spaced data points. The program allows interactive, real time curve fitting by the operator and/or objective curve fitting using a non-linear least squares fitting routine, or a combination of the two. The program is more robust in a micro-computer environment than subtractive or generalized non-linear least squares methods and faster than pattern searching methods. The program has been used over the last 4 years in my laboratory by a number of personnel without any previous experience of computers. It is written in BASIC and runs on a Hewlett-Packard series 200 model 16S (9816) computer or on a Hewlett-Packard model 9845S computer. The program is structured to allow use of any underlying curve shape with 3 parameters per component.