Laparoscopy in the second and third trimesters of pregnancy for abdominal surgical emergencies.

OBJECTIVE: To assess complications and outcomes of pregnancies following laparoscopic abdominal surgery during the second and third trimesters of pregnancy. MATERIAL AND METHODS: Retrospective single-center study of 23 cases of laparoscopic surgery in the second or third trimesters of pregnancy between January 2005 and May 2016. RESULTS: The laparoscopies were performed between 15 and 33 weeks of gestation, a mean of 23 weeks+2 days, with 6 cases in the 3rd trimester. The operations were: 11 cholecystectomies, 6 appendectomies, 1 intestinal occlusion (volvulus on a gastric band), 3 adnexal torsions, 1 ovarian cyst and 1 paratubal cyst with torsion. No secondary laparotomy was required. The postoperative courses were favorable in most cases. However, 3 appendectomies were complicated, one by chorioamnionitis and miscarriage at 20½ weeks of gestation and 2 by right iliac fossa abscesses requiring percutaneous radiological drainage, one of these women delivered a healthy term baby and the other had chorioamnionitis and preterm delivery at 34 weeks, followed by neonatal death. CONCLUSION: Laparoscopy can be safely performed for surgical indications in the second and third trimesters of pregnancy. In case of abdominal symptoms, a timely diagnosis is required to decide whether or not to operate and imaging should not be withheld particularly in case of suspected appendicitis which has a high risk of complications.

An analysis of hospital productivity and product change.

We developed a model to measure the contribution of changes in length-of-stay, service intensity, and productivity to the unusually low rate of growth in hospital costs per discharge in recent years. From 1992 through 1996 declining length-of-stay explained 97 percent of the decrease in real costs per discharge. Much of the drop was probably caused by care shifted from inpatient to postacute settings. Although complete data for our model are unavailable beyond that point, we cite several "leading indicators" that suggest that length-of-stay declines have played a smaller role in the continued low cost growth of 1997 and 1998 and that productivity may have risen sharply.

Cost-effective medicine: the financial impact that practice guidelines have on outpatient hospital charges in the emergency department.

The objective of this study was to determine whether specific cost-effective guidelines based on a patient's chief complaint can significantly reduce outpatient hospital charges in the ED. A prospective randomized single-blinded clinical trial was conducted in an urban community hospital ED with a 14,000 annual census. The first phase of the study involved preintervention data collection. The second phase focused on development, physician approval, and implementation of 23 specific cost-effective guidelines, as well as general recommendations for diagnostic tests. The third phase involved postintervention data collection. Results showed that the total outpatient hospital charge decreased by 28% per patient. The laboratory hospital charge decreased by 46% per patient. The radiology hospital charge decreased by 20% per patient. The hospital supply charge and pharmacy charge decreased by 31% per patient and 11% per patient, respectively. In conclusion, cost-effective medicine practiced with specific guidelines, based on a patient's chief complaint, significantly reduces unnecessary diagnostic tests and medical treatments ordered by emergency physicians.

Hospital-physician arrangements and hospital financial performance.

OBJECTIVES: The introduction of the Medicare Prospective Payment System and the more recent rise of managed care plans have greatly increased the importance of effective hospital financial management. Because physicians play a central role in directing hospital resource use, policies to influence physician behavior and to align physician and hospital interests more effectively are being advocated increasingly. This article evaluates the effect of nine strategies to facilitate physician involvement and integration into the hospital on hospital financial performance. METHODS: Data came primarily from the Prospective Payment Assessment Commission's hospital-physician relations survey of 1,485 hospitals and the Medicare Cost Reports. Both ordinary least squares and first differencing models were used to evaluate the effect of physician integration on hospital financial performance. RESULTS: Hospitals with lower margins and higher costs were more likely to have implemented strategies to integrate physicians and to modify physician behavior than their counterparts. Analysis using first differencing models indicated that making department heads responsible for the profits and losses had a significant positive effect on margins, whereas including medical staff on the hospital's board and offering physicians management services had a significant negative impact on average Medicare costs. In addition the number of strategies implemented was associated positively with financial performance. The paper also emphasizes the importance of model specification in evaluations of hospital-physician arrangements. CONCLUSIONS: Changes in hospital-physician relations may have been one reason why hospitals have been relatively successful at containing costs and retaining profitability in recent years. More research needs to be done on which specific arrangements affect hospital financial performance, as well as their effect on the quality of patient care.

Estimating increases in outpatient dialysis costs resulting from scientific and technological advancement.

The Medicare program's base payment rate for outpatient dialysis services has never been adjusted for the effects of inflation, productivity changes, or scientific and technological advancement on the costs of treating patients with end-stage renal disease. In recognition of this, Congress asked the Prospective Payment Assessment Commission to annually recommend an adjustment to Medicare's base payment rate to dialysis facilities. One component of this adjustment addresses the cost-increasing effects of technological change--the scientific and technological advances (S&TA) component. The S&TA component is intended to encourage dialysis facilities to adopt technologies that, when applied appropriately, enhance the quality of patient care, even though they may also increase costs. We found the appropriate increase to the composite payment rate for Medicare outpatient dialysis services in fiscal year 1995 to vary from 0.18% to 2.18%. These estimates depend on whether one accounts for the lack of previous adjustments to the composite rate. Mathematically, the S&TA adjustment also depends on whether one considers the likelihood of missing some dialysis sessions because of illness or hospitalization. The S&TA estimates also allow for differences in the incremental costs of technological change that are based on the varying advice of experts in the dialysis industry. The major contributors to the cost of technological change in dialysis services are the use of twin-bag disconnect peritoneal dialysis systems, automated peritoneal dialysis cyclers, and the new generation of hemodialysis machines currently on the market. Factors beyond the control of dialysis facility personnel that influence the cost of patient care should be considered when payment rates are set, and those rates should be updated as market conditions change. The S&TA adjustment is one example of how the composite rate payment system for outpatient dialysis services can be modified to provide appropriate incentives for producing high-quality care efficiently.

Protease inhibitor display M13 phage: selection of high-affinity neutrophil elastase inhibitors.

We report display of the complete protease inhibitor (Kunitz) domain, BPTI, on the surface of bacteriophage M13 as a fusion to the gene III product. Phage that display BPTI bind specifically to anti-BPTI antibodies, trypsin and anhydrotrypsin. A point mutation of BPTI [Lys15-->Leu(K15L)] alters the binding specificity of fusion phage such that a human neutrophil elastase-binding phenotype is conferred while a trypsin-binding phenotype is eliminated. Phage were eluted from an immobilized protease with step gradients of decreasing pH. Phage that display Kunitz domains having higher affinity for the immobilized protease exhibit characteristic pH elution phenotypes, indicating that bound display phage can be selectively recovered from an affinity matrix. Utilization of this technology should enable the selection of remodeled protease inhibitors exhibiting novel binding specificities.

Directed evolution of a protein: selection of potent neutrophil elastase inhibitors displayed on M13 fusion phage.

Inhibitors of human neutrophil elastase were engineered by designing and producing a library of phage-displayed protease inhibitory domains derived from wild-type bovine pancreatic trypsin inhibitor and fractionating the library for binding to the target protease. The affinity of one of the engineered variants for human neutrophil elastase (Kd = 1.0 pM) is 3.6 x 10(6)-fold higher than that of the parental protein and exceeds the highest affinity reported for any reversible human neutrophil elastase inhibitor by 50-fold. Thus the display phage method has allowed us to obtain protein derivatives that exhibit greatly increased affinity for a predetermined target. The technology can be applied to design high-affinity proteins for a wide variety of target molecules.

Design, construction and function of a multicopy display vector using fusions to the major coat protein of bacteriophage M13.

Incorporation of numerous copies of a heterologous protein (bovine pancreatic trypsin inhibitor; BPTI) fused to the mature major coat protein (gene VIII product; VIII) of bacteriophage M13 has been demonstrated. Optimization of the promoter, signal peptide and host bacterial strain allowed for the construction of a working vector consisting of the M13 genome, into which was cloned a synthetic gene composed of a lac (or tac) promoter, and sequences encoding the bacterial alkaline phosphatase signal peptide, mature BPTI and the mature coat protein. Processing of the BPTI-VIII fusion protein and its incorporation into the bacteriophage were found to be maximal in a host bacterial strain containing a prlA/secY mutation. Functional protein is displayed on the surface of M13 phage, as judged by specific interactions with antiserum, anhydrotrypsin, and trypsin. Such display vectors can be used for epitope mapping, production of artificial vaccines and the screening of diverse libraries of proteins or peptides having affinity for a chosen ligand. The VIII display phage system has practical advantages over the III display phage system in that many more copies of the fusion protein can be displayed per phage particle and the presence of the VII fusion protein has little or no effect on the infectivity of the resulting bacteriophage.

Streptomyces genes involved in biosynthesis of the peptide antibiotic valinomycin.

We have identified genes from Streptomyces levoris A-9 involved in the biosynthesis of the peptide antibiotic valinomycin. Two segments of chromosomal DNA were recovered from genomic libraries, constructed by using the low-copy-number plasmid pIJ922, by complementation of valinomycin-deficient (vlm) mutants of S. levoris A-9. One set of plasmids restored valinomycin production to only one mutant, that carrying vlm-1, whereas a second set of plasmids restored productivity to seven vlm mutants, those carrying vlm-2 through vlm-8. Additional complementation studies using subcloned restriction enzyme fragments showed that the vlm-1+ gene was contained within a 2.5-kilobase (kb) DNA region, whereas alleles vlm-2+ through vlm-8+ were contained in a 12-kb region, representing at least three genes. Physical mapping experiments based on the isolation of cosmid clones showed that the two vlm loci were 50 to 70 kb apart. Southern hybridization experiments demonstrated that the vlm-2+ gene cluster was highly conserved among other valinomycin-producing Streptomyces strains, whereas the vlm-1+ gene was ubiquitous among Streptomyces species tested. Increasing the copy number of the vlm-2+ gene cluster in S. levoris A-9 by the introduction of low-copy-number recombinant plasmids resulted in a concomitant increase in the level of valinomycin production.

The first 3 years of Medicare prospective payment: an overview.

This article provides a synopsis of the available evidence on the impact of the Medicare prospective payment system (PPS) for hospitals over the first 3 years of its implementation. The impact of PPS on hospitals, Medicare beneficiaries, post-hospital care, other payers for inpatient hospital services, other health care providers, and Medicare program operations and expenditures is examined.

The rho-115 mutation in transcription termination factor rho affects its primary polynucleotide binding site.

We have investigated the effect of the rho-115 mutation on the catalytic properties of the Escherichia coli termination protein, rho. Comparison of the primary and secondary polynucleotide binding sites activities reveals dramatic differences between the mutant and wild-type molecules. Wild-type rho must bind single-stranded polynucleotides to activate its nucleotide triphosphatase (NTPase) activity, and either poly(C), or poly(dC) plus oligo(C), will suffice. In contrast, attempted activation of the rho-115 NTPase with poly(C) in the presence of poly(dC) showed the latter to be a potent inhibitor. Inclusion of small oligonucleotides such as oligo(C) in the activation assay does not inhibit the poly(C)-induced NTPase reaction of either wild-type rho or rho-115. This would indicate, in the two polynucleotide binding site model for rho proposed by Richardson (Richardson, J.P. (1982) J. Biol. Chem. 251, 5760-5766), that the mutation in rho-115 affects the primary polynucleotide binding site. Transcription termination in vitro at the rho-dependent site trp t' showed dramatically reduced termination with rho-115 protein compared to wild-type rho. In the presence of rho-115, the transcript is longer and termination occurs over a narrower range of nucleotides than with wild-type rho. This suggests that the primary polynucleotide binding site is important not only for efficient termination of transcription but may also be involved in determining the terminal end point of the transcript itself.

Impact of the Medicare prospective payment system for hospitals.

This article describes some of the available evidence on the impact of the Medicare prospective payment system (PPS) for hospitals during its first year, on hospitals, other payers for inpatient hospital services, other providers of health care, and Medicare beneficiaries. In addition, because the impetus for the enactment of the new system stemmed from concern over the financial status of the Medicare program, the first-year impact of PPS on Medicare program expenditures is also described.

Purification of RNA polymerase and transcription-termination factor Rho from Erwinia carotovora.

Erwinia carotovora RNA polymerase consists of the holoenzyme structure sigma 2 beta beta' sigma as found in Escherichia coli and other bacteria. E. carotovora RNA polymerase can synthesize RNA using lambda, T7 of T4 DNA as templates; however, it is two times less active on these templates and is more temperature-sensitive than the E. coli enzyme. The alpha subunit of the E.. carotovora enzyme is lower in molecular mass than its E. coli counterpart. The sigma factors from E. coli and E. carotovora are similar in size and in their ability to stimulate RNA synthesis by core enzyme on DNA templates such as T7 DNA. An additional protein of 115 000 Da molecular mass, termed gamma, is found associated with E. carotovora RNA polymerase. The gamma protein is tightly associated with the polymerase subunits as it is not dissociated by gel filtration in buffer containing 0.5 M NaCl. It can be purified by passing the Agarose 1.5 m enzyme through coupled Bio-Rex 70 and DEAE-cellulose columns. The gamma-protein, when present in excess over the sigma subunit, inhibits holoenzyme activity on T7 DNA but not on poly[d(A-T)]and may thus interfere with sigma activity. The gamma protein by itself cannot transcribe T7 DNA or poly[d(A-T)], nor does it stimulate core enzyme activity on T7 DNA. E. carotovora rho has a subunit molecular mass of 48 000 Da and exhibits RNA-dependent phosphohydrolysis of adenosine ribonucleoside triphosphate. E. coli and E. carotovora rho are indistinguishable immunologically, as total fusion of precipitin bands is observed. E. carotovora rho elutes from a phosphocellulose column at a salt concentration of about 0.21 M KCl, compared to that of 0.29 M KCl for E. coli rho. The poly(C)-dependent ATPase activity of E. carotovora rho is more-temperature sensitive and is six to ten times less active than that of E. coli rho. E. carotovora rho is capable of terminating RNA transcripts, as indicated by a decrease in RNA synthesis using lambda or T7 DNA as template and E. carotovora or E. coli polymerase as the transcribing-enzyme.

Location of the rho gene and characterization of lambda ilv-gal derivatives of lambda ilv-rho bacteriophage.

The location of the rho gene and its position relative to the ilv genes of Escherichia coli K-12 was analyzed using genetic criteria, restriction enzyme cleavage, and maxicell analysis. Plasmids were constructed with deletions of the rho gene introduced in vitro, and lambda ilv-gal derivatives of lambda ilv-rho bacteriophage were isolated by recombination in vivo. A HindIII restriction fragment of 8 kilobases (kb) previously shown to contain at least part of the rho gene (Gray et al. 1981) was cloned into plasmid pMC81. This vector has transcription stop sites that present read-through expression of cloned genes from either direction, and cloning sites upstream of the lacZ gene coding for beta-galactosidase. The position of the rho gene and flanking sequences required for its expression were further localized to a region of approximately 2 kb by introducing deletions using restriction enzyme treatment of these plasmids. A promoter in the rho region was found to direct beta-galactosidase synthesis in these plasmid derivatives. Derivatives of lambda ilv-rho phage were isolated in vivo by pyrophosphate chelation selection for phage with reduced genome size. Restriction enzyme analysis of twelve of these derivatives revealed an unexpected bias towards phage recombinants as opposed to simple internal deletions.

Autoregulation of the rho gene of Escherichia coli K-12.

It has previously been proposed, based on indirect evidence, that the Rho protein may control the expression of the rho gene. Using an in vitro system for the transcription and translation of the rho gene cloned into plasmid pBR322, we tested this hypothesis directly by monitoring the effect in vitro of excess or limiting Rho protein. The addition of purified Rho protein suppresses Rho synthesis in vitro. The addition of antibody to Rho specifically stimulates Rho synthesis in vitro. The stimulation of Rho factor synthesis by antibody to Rho is reversed by Rho protein. Rho factor purified from a strain with a mutationally altered rho gene (rho-115) does not suppress Rho synthesis in vitro. These results provide convincing evidence that the rho gene is subject to autoregulation.

Pyrophosphate inhibition of rho ATPase: a mechanism of coupling to RNA polymerase activity.

The effects of pyrophosphate on RNA binding and ATPase activities of Escherichia coli transcription termination factor rho have been studied. Mutant rho-115 protein has a temperature-sensitive RNA-dependent ATPase activity due to the thermolability of binding to RNA [Kent, R.B. & Guterman, S.K. (1981) Fed. Proc. Fed. Am. Soc. Exp. Biol. 40, 1765 (abstr.)]. The presence of either ATP or pyrophosphate at comparable concentrations stabilizes the binary complex of rho and poly(C) at high temperature. ADP at 8-fold greater concentration also stabilizes the mutant rho-RNA binary complex. Pyrophosphate is a noncompetitive inhibitor (Ki = 0.07 mM) of rho poly(C)-dependent ATPase, an activity that is required for rho-mediated termination. These results suggest the existence of a regulatory site on the rho molecule. We suggest that rho NTPase is regulated by RNA polymerase (EC 2.7.7.6) so that during transcription elongation the RNA polymerase competes successfully with rho for substrates and inhibits rho NTPase with product pyrophosphate. Further, RNA polymerase pausing may result in reduced pyrophosphate and increased NTP concentrations, allowing rho NTPase to function.

Polarity in the glnA operon: suppression of the reg- phenotype by rho mutations.

To determine the ability of mutations in glnA, the gene for glutamine synthetase (GS), to regulate nitrogen assimilatory enzymes, we assayed histidase and GS in 34 glnA (Gln(-)) strains. Twenty-five glnA mutants were RegC, synthesizing high levels of histidase regardless of the availability of nitrogen, and nine were Reg(-), synthesizing low levels of histidase in medium containing either limiting or excess ammonia. rho mutations were introduced into strains containing glnA point mutations or insertions in glnA, glnL, glnG, or glnF. The Reg(-) phenotype of strains with glnA point mutations, but not those with glnA or glnF insertions, was altered by the presence of rho, suggesting that glnA (Reg(-)) mutations are polar and exert their phenotype by decreasing expression of glnL and glnG. Consistent with this view, no GS protein was detected by two-dimensional gel electrophoresis in glnA (Reg(-)) rho(+) or glnA (Reg(-)) rho double mutants, whereas GS protein was detected in cells of 10 of 11 glnA (RegC) strains. Since glnA (Reg(-)) rho double mutants synthesize constitutive levels of histidase, GS protein is not necessary for full expression of histidase. Mu d1 insertions in glnL, but not those in glnG, responded to the presence of a rho allele, presumably owing to elevated transcription into glnG from the Mu d1 prophage. Our results suggest that glnA (Reg(-)) alleles are polar mutations, and a rho-dependent termination site down-stream is postulated as the basis for the polar phenomenon. The data also indicate that, under some circumstances, a significant portion of glnL and glnG transcription is initiated at the glnA promoter.