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Biomedical subjects

J P Murphy

Publications and source records attributed to J P Murphy.

At least 19 recordsLinked to original sources
PubMed1992

Follicular lymphoma with t(8;14)(q24;q32): a distinct clinical and molecular subset of t(8;14)-bearing lymphomas.

The presence of the translocation t(8;14)(q24;q32) has not been well described in follicular lymphoma (FL). In a consecutive series of 278 karyotypically abnormal non-Hodgkin's lymphomas (NHL), six patients with FL showing a t(8;14) without a t(14;18)(q32;q21) were identified. They ranged in age from 45 to 73 years. The cell type was mixed in four patients, small-cleaved in one, and large-cleaved in one; four cases also contained diffuse areas. All cases tested displayed monoclonal surface Ig. The clinical courses were consistent with the histologic subtypes, being less aggressive than other t(8;14)-bearing NHL. In five cases, frozen tissue was available for Southern blotting. The BCL2 gene showed a germline configuration when studied with the MBR, MCR, and 5' cDNA probes. The MYC gene also appeared unrearranged using an exon-1 probe with EcoRI or HindIII digestion. Analysis of the Ig heavy chain (IgH) gene with a JH region probe and BamHI or EcoRI digestion showed only one rearranged band in all cases, indicating that the 14q32 breakpoint did not lie in either the J or switch-mu (SM) regions. In four cases, the exon-1/intron-1 border of the MYC gene, a target area for point mutations in cases of t(8;14) that do not display rearrangements of the MYC gene, was enzymatically amplified and sequenced; no point mutations were identified. The indolent behavior of our six cases, and the finding that the molecular structure of the t(8;14) in these cases does not follow the pattern of breakpoint sites and point mutations defined in other histologic subtypes of NHL with this translocation, suggest that the t(8;14) in these cases is cytogenetically and molecularly distinct from the t(8;14) seen in high-grade NHLs, and is relatively ineffectual in terms of MYC deregulation, or that other genetic elements at these chromosomal sites may be involved. Further analysis of these tumors may provide insights into MYC deregulation and BCL2-independent FL.

Aged↗
PubMed1992

Escalator injuries.

A series of 50 patients suffering injuries from falls on escalators attended our accident and emergency department. Of these, 8 (16 per cent) required admission. Alcohol was a factor in 11/15 (74 per cent) of men but in only 2/34 (6 per cent) of women. Of the injuries, 32 (64 per cent) were associated with the practice of walking on the escalator, while only 12 (24 per cent) were associated with stepping onto or off the escalator. Discouraging the practice of walking on one side of the escalator might help to minimize injuries.

Accidents↗
PubMed1992

Congenital diaphragmatic hernia presenting due to gastrointestinal complications.

Congenital diaphragmatic hernia (CDH) presents beyond the first hours of life in 10% to 20% of cases. Presenting symptoms may be quite nonspecific, and are often gastrointestinal rather than respiratory in origin. We have recently had experience with five such cases, one in a newborn and four in older children. All presented with symptoms related to gastrointestinal complications of their diaphragmatic defect. In the newborn, gastric perforation had occurred, a complication of this anomaly not previously reported. The chest radiograph showed loops of bowel in the chest in all cases, allowing correct preoperative diagnoses. Urgent operative intervention was undertaken in each case with good results and no long-term morbidity. The risk of intestinal strangulation in the late-presenting CDH patient warrants emergent surgical management, which should be rewarded by uniform survival with few complications. Although preoperative stabilization may decrease the severity of pulmonary vasospasm in the newborn with respiratory failure, delay may increase the risk of bowel infarction in the older child presenting with gastrointestinal symptoms.

Child, Preschool↗
PubMed1992

Amplified expression and large-scale purification of protein G'.

PCR was used to isolate the gene fragment coding for Protein G' (SpG'), a truncated bacterial cell surface protein from Streptococcus G148 which binds to the Fc region of IgG and expressed in E. coli [Goward et al. (1990) Biochem. J. 267: 171-177]. The PCR primer was designed to change the TTG initiation triplet to ATG and to incorporate it into an NdeI restriction site (CATATG), allowing the gene to be cloned in frame into an NdeI restriction site immediately downstream of a trp promoter. Expression of SpG' was estimated as about 30% total soluble cell protein which compares very favourably to the less than 1% total soluble cell protein obtained from the original system [Goward, et al. (1990) Biochem. J. 267: 171-177]. Homogeneous SpG' was recovered by a single anion-exchange chromatography step on Q-Sepharose FF in a process which avoided use of an affinity adsorbent. Even though SpG' consists of almost identical repetitive domains from amino acid sequence analysis, different proteolytic sensitivity of each domain was observed indicating their structural dissimilarity.

Amino Acid Sequence↗
PubMed1991

Sequential 1H NMR assignments and secondary structure of an IgG-binding domain from protein G.

Protein G is a member of a class of cell surface bacterial proteins from Streptococcus that bind IgG with high affinity. A fragment of molecular mass 6988, which retains IgG-binding activity, has been generated by proteolytic digestion and analyzed by 1H NMR. Two-dimensional DQF-COSY, TOCSY, and NOESY spectra have been employed to assign the 1H NMR spectrum of the peptide. Elements of regular secondary structure have been identified by using nuclear Overhauser enhancement, coupling constant, and amide proton exchange data. The secondary structure consists of a central alpha-helix (Ala28-Val44), flanked by two portions of beta-sheet (Val5-Val26 and Asp45-Lys62). This is a fundamentally different arrangement of secondary structure from that of protein A, which is made up of three consecutive alpha-helices in free solution (Torigoe et al., 1990). We conclude that the molecular mechanisms underlying the association of protein A and protein G with IgG are different.

Amino Acid Sequence↗
PubMed1991

The secondary structure of protein G', a robust molecule.

The secondary structure of recombinant streptococcal Protein G' was predicted and compared with spectropolarimetric data. The predicted secondary structure consisted of 37 +/- 4% alpha-helix and 30 +/- 5% beta-sheet, whereas the values obtained from c.d. data were 29 +/- 2% alpha-helix and 41 +/- 3% beta-sheet. An alpha-helix-beta-sheet/turn-alpha-helix motif is conjectured to comprise the Fc-binding unit. The c.d. spectra in the near u.v. and far u.v. show that the Protein G' molecule is stable to heating at 100 degrees C and to extremes of pH (pH 1.5 to 11.0). The protein retained biological activity at these extremes. The molecule uncoils above pH 11.5 in a time-dependent fashion. Unfolding of the molecule in guanidinium chloride was monitored by c.d. and fluorescence emission; 3 M-guanidinium chloride was required to unfold the protein by 50%. The protein was completely unfolded in 5.5 M-guanidinium chloride and fully refolded with restoration of activity after removal of guanidinium chloride.

Amino Acid Sequence↗
PubMed1990

Expression and purification of a truncated recombinant streptococcal protein G.

The gene for Protein G from Streptococcus strain G148 was cloned and expressed in Escherichia coli. The regions on the gene corresponding to the albumin-binding domains and the Fab-binding region were then deleted by site-directed mutagenesis. The translation of regions corresponding to the cell-wall- and membrane-binding domains was prevented by introduction of stop codons upstream of these domains. This recombinant DNA sequence codes for a protein (G') that contains repetitive regions and that binds only the Fc portion of IgG, analogously to Protein A. Translation of the sequence produces a protein with an Mr of about 20,000. The nucleotide sequence differs from those published previously [Guss, Eliasson, Olsson, Uhlén, Frej, Jornvall, Flock & Lindberg (1986) EMBO J. 5, 1567-1575; Olsson, Eliasson, Guss, Nilsson, Hellman, Lindberg & Uhlén (1987) Eur. J. Biochem. 168, 319-324]. The protein can be substantially purified on a large scale by chromatography on IgG-Sepharose 4B. Homogeneous Protein G' can be prepared by anion-exchange f.p.l.c. on Mono Q HR. This Protein G' has a pI of 4.19 and SDS/PAGE gives an apparent anomalous Mr of 35,000.

Amino Acid Sequence↗
PubMed1990

Rectal prolapse: 17-year experience with the posterior repair and suspension.

Using a posterior repair and rectal suspension procedure for those patients who need surgical treatment of rectal prolapse, we have treated 46 patients over a period of 17 years at Children's Mercy Hospital in Kansas City, MO. One patient with caudal dysgenesis died of multiple congenital anomalies following two unsuccessful attempts at posterior repair and suspension. Four patients developed a recurrence afterwards, which was found to be due to sigmoid intussusception and, presumably, had played a major part in their original prolapse. Two of these required resection, one from the transanal approach and one from the transabdominal approach. One resolved spontaneously and another is as yet unresolved. Three patients had minor mucosal prolapse that was transient and two patients had extrusion of silk sutures but continued to have a very satisfactory result. Overall, 42 patients had satisfactory resolution of their rectal prolapse. Three of the four patients who had unsatisfactory results had associated anomalies that contributed to their poor outcome.

Child↗
PubMed1989

Ras oncogene mutation in multiple myeloma.

The frequency of ras (H-, K-, and N-ras) and c-myc oncogenes was investigated in multiple myeloma (MM). By means of the polymerase chain reaction (PCR)/oligonucleotide hybridization method, DNA from 56 tumor biopsies was analyzed for the presence of activating mutations involving codons 12 and 61 of the H-, K-, and N-ras genes and codon 13 of the N-ras gene. Mutations, involving the N- or K-ras genes, were detected in 18 of 56 (32%) cases of which 12/43 (27%) were at diagnosis and 6/13 (46%) were after treatment. In some cases, multiple mutations affecting different ras alleles were detected. Direct nucleotide sequence analysis of PCR products indicated that a more heterogeneous nature of the base pair changes than previously shown for other tumors along with a preferential involvement of N-ras codon 61. The heterogeneity of MM cases with respect to the presence of ras oncogenes prompted an analysis of possible correlations with different clinico-pathologic characteristics of MM from which a correlation between the presence of ras oncogenes and a partial or complete lack of response to therapy emerged. The frequency of activating rearrangements or mutations of the c-myc gene were studied by Southern blot analysis and PCR sequencing, respectively. However, contrary to previous reports involving mostly MM cell lines, no structural alterations of the c-myc gene were found. These results indicate that ras, but not c-myc, oncogenes are activated in vivo in MM cells, representing the first oncogene alteration that has been associated at appreciable frequency with this type of malignancy. While the mechanism of occurrence and biological role of ras activation in MM remains to be elucidated, the preliminary correlations observed in this study between the presence of ras oncogenes and poor therapeutic response suggest that further investigations of the possible prognostic significance of these alterations are necessary.

Base Sequence↗
PubMed1989

Continent urinary reconstruction in ischiopagus tripus conjoined twins.

The principle of the continent urinary reservoir involves use of a variety of intestinal segments providing continence, a mechanism of antireflux and a catheterizable stoma in either an abdominal or pelvic location. This concept was used to create a continent urinary reservoir in a 3-year-old former ischiopagus tripus conjoined twin. The solitary renal unit had been drained into a hydrocolpos with an antirefluxing ureteral reimplantation at separation. The patient remained incontinent through the urogenital sinus. At subsequent reconstruction the posterior aspect of the hydrocolpos was tubularized as a vagina, while the remainder of the hydrocolpos was augmented with ileum to create a urinary reservoir. The conduit was constructed with imbricated ileum, in which myectomy had been performed to allow for easier imbrication and tubularization for a narrow neourethra. A second set of conjoined twins have been separated similarly. Of these twins 1 will undergo creation of a continent urinary reservoir in an identical fashion. This represents the first report of the use of hydrocolpos in the creation of a urinary reservoir.

Child, Preschool↗
PubMed1988

Overproduction of protein kinase C causes disordered growth control in rat fibroblasts.

We have generated a series of rat fibroblast cell lines that stably overexpress a full-length cDNA encoding the beta 1 form of protein kinase C (PKC). These cell lines contain a 20- to 53-fold increase in PKC activity and exhibit dramatically enhanced morphologic changes following exposure to the tumor promoter 12-O-tetradecanoyl phorbol-13-acetate (TPA). They grow to a high saturation density in monolayer cultures and, when maintained at postconfluence, develop small, dense foci. In contrast to control cells, which display complete anchorage dependence, PKC-overproducing cells form small colonies in soft agar in the absence of TPA and large colonies in the presence of TPA. Thus, the mere overproduction of a single form of PKC is sufficient to confer multiple growth abnormalities in rat fibroblasts. These results provide direct evidence that PKC plays a critical role in growth control and that it mediates several of the cellular effects of the phorbol ester tumor promoters. They also suggest that the activation of PKC may be of central importance in the process of multistage carcinogenesis.

Amino Acid Sequence↗
PubMed1988

Hepatic hemangioma: resection using hypothermic circulatory arrest in the newborn.

Hepatic hemangioma (hemangioendothelioma) is a benign vascular tumor with significant morbidity and mortality. Two newborn infants underwent partial hepatectomy for treatment of massive hepatic hemangioma, one with atrial septal defect and secondary congestive heart failure, and the other with symptomatic marked hepatomegaly. Cardiopulmonary bypass with induced deep hypothermic circulatory arrest was used in both patients in order to facilitate resection and prevent sudden massive blood loss. One infant also had closure of an atrial septal defect in addition to the partial hepatectomy. Both infants survived without complication and showed complete resolution of preoperative symptoms. This technique coupled with the use of fibrin glue is of value in infants with large hepatic hemangiomata and cardiac decompensation in the neonatal period.

Cardiopulmonary Bypass↗
PubMed1988

Histology of the intestine in human gastroschisis--relationship to intestinal malfunction: dissolution of the "peel" and its ultrastructural characteristics.

There are conflicting views on the pathogenesis of the intestinal malfunction seen in infants with gastroschisis. It has been variously ascribed to abnormalities of ganglion cells and smooth muscle elements, intestinal ischemia, and the "peel" which invests the serosa of the intestine. Review of the clinical and experimental literature showed only limited information on the histology of the eviscerated human intestine. In order to add to this data base, and to further investigate the pathogenesis of the intestinal malfunction from a histologic standpoint, we reviewed surgical and autopsy material from our experience with 105 neonates with gastroschisis. Ten specimens were satisfactory for evaluation from a standpoint of tissue integrity. The specific mural components of mucosa, submucosa, muscularis, and ganglion cells were examined and found to be either normal, or to show nonspecific abnormalities that varied from case to case, and were related mostly to intestinal infarction due to compromise of the gut at the site of the gastroschisis defect. In six patients, this progressed to atresia formation. The most consistent abnormalities were found in the serosal layer with its peel. Using special stains, the peel was found to be composed largely of fibrin and collagen. Based on this study, we feel that edema and ischemic changes, though often present, are much less prominent than the peel, as the leading histologic abnormality of the intestine of gastroschisis. Squamous epithelial cells were seen in the peel in four cases, suggesting that the peel had been "appliqued" onto the serosa of the herniated fetal gut.

Abdominal Muscles↗
PubMed1987

Amino acid sequence of the L-lactate dehydrogenase of Bacillus caldotenax deduced from the nucleotide sequence of the cloned gene.

The Bacillus caldotenax L-lactate dehydrogenase gene (lct) has been cloned into Escherichia coli, using the Bacillus stearothermophilus lct gene as a hybridisation probe, and its complete nucleotide sequence determined. The lct structural gene consists of an open reading frame of 951 base pairs commencing with an ATG start codon and followed by a TAA stop codon. Upstream of the gene are putative transcriptional promoter -35 and -10 regions; a ribosome binding site with a predicted delta G of -66.9 kJ/mol is also present six base pairs upstream of the ATG start codon. The B. caldotenax lct gene is highly homologous to the B. stearothermophilus lct gene displaying a DNA sequence homology of 89.7%. Examination of the DNA sequence 3' of the lct gene revealed the presence of two further open reading frames. This suggests that the lct gene may be the first gene of an operon. The deduced amino acid sequence of the L-lactate dehydrogenase (LDH) from B. caldotenax predicted a protein of 317 amino acid residues; comparison with the B. stearothermophilus enzyme revealed only 30 amino acid differences between the two enzymes; thus the enzymes are 90.4% homologous. These amino acid differences must account for the different thermostabilities of the two enzymes. The B. caldotenax lct gene was efficiently expressed in E. coli and the original lct-containing plasmid construct isolated (pKD1) induced the synthesis of LDH at a level of 4.5% of the E. coli soluble cell protein whilst a SmaI subfragment of this clone, (pKD2) produced LDH at a level of 6.9% of the E. coli soluble cell protein. LDH isolated from E. coli cells had the same thermal stability properties as LDH isolated from B. caldotenax cells.

Amino Acid Sequence↗
PubMed1986

Retinoic acid inhibits junctional communication between animal cells.

Retinoic acid inhibits junctional communication between a variety of vertebrate cell types in culture. It reduces the intercellular transfer of 3H-nucleotides between Syrian hamster kidney fibroblasts (BHK 21/13), Chinese hamster lung fibroblasts (V79), rat liver epithelial cells (BRL), Swiss mouse embryo fibroblasts (3T3), rainbow trout gonadal fibroblasts (RTG2) and Xenopus embryo fibroblasts (Xen). It also reduces metabolic cooperation between hypoxanthine-guanine phosphoribosyl transferase deficient mutant and wild-type BHK cells. The inhibition is rapid (intercellular transfer of iontophoretically injected Lucifer Yellow CH between BRL cells is completely blocked after the cells have been exposed to 10(-4) M retinoic acid for 5 min), and is fully reversed when the drug is removed. Based on these results and the observation that the amount of gap junctional protein isolated from cells grown in the presence of retinoic acid for 1 h is the same and after 24 h is increased (1.3- to 3.1-fold) compared with the amount isolated from untreated cells, we suggest that the inhibitory effect is mediated by the reversible closure of junctional channels.

1-Octanol↗