Cost-benefit analysis and short-term outcomes after implementing an ERAS protocol for colorectal surgery: a propensity score-matched analysis.

BACKGROUND: Enhanced Recovery After Surgery (ERAS) has become increasingly popular in the post-operative management of abdominal surgery. Published data suggest that patients on ERAS protocols have fewer minor and major complications, and highlight a reduction in medical morbidity (such as urinary and respiratory infections). Limited data is available on surgical complications. The aim of the study was to evaluate the impact of the ERAS protocol on post-operative complications and length of hospital stay. Furthermore, we aimed to determine the impact of this protocol on cost-effectiveness. MATERIAL AND METHODS: From January 2016 to December 2022, 532 colectomies for colorectal cancer (CRC) were performed. A prospective observational study was conducted in a tertiary hospital on the cohort of patients, aged 18 years and older, operated on for non-urgent colorectal cancer. The impact on post-operative complications, hospital stay and economic impact was analysed in two groups: patients managed under ERAS and non-ERAS protocol. A propensity score-matching analysis was performed between the two groups. RESULTS: After propensity score matching 1:1, each cohort included 71 patients, and clinicopathological characteristics were well balanced in terms of tumour type, surgical technique and surgical approach. ERAS patients experienced fewer infectious complications and a shorter postoperative stay (p < 0.001). In particular, they had an 8.5% reduction in anastomotic dehiscence (p = 0.012) and surgical wound infections (p = 0.029). After analysis of medical complications, no statistically significant differences were identified in urinary tract infections, pneumonia, gastrointestinal bleeding or sepsis. ERAS protocol was more efficient and cost-effective than the control group, with an overall savings of 37,673.44€. CONCLUSIONS: The implementation of an enhanced recovery protocol for elective colorectal surgery in a tertiary hospital was cost-effective and associated with a reduction in post-operative complications, especially infectious complications.

Incidence and risk factors for incisional hernia after temporary loop ileostomy closure: choosing candidates for prophylactic mesh placement.

OBJECTIVES: The primary aim of this study was to identify the incisional hernia rate after temporary loop ileostomy closure. Secondary outcomes were determining the risk factors linked to this incisional hernia, which could improve the patient selection for mesh prophylaxis. METHODS: Retrospective cohort study of all consecutive patients with temporary loop ileostomy reversal through a peristomal incision from 1st January 2011 to 1st January 2017 at our centre. Data were extracted from electronic clinical records: baseline patient characteristics, surgical factors and postoperative events. CT scans performed during follow-up were precisely analysed. Survival analysis was applied to identify risk factors for hernia development. RESULTS: 129 patients were analysed of whom 15 (11.6%) developed an incisional hernia at previous ileostomy site. The median time for incisional hernia development was 11 months (IQR = 7-21) and the median follow-up time was 37 months (IQR = 22-57). The identified patient risk factors for hernia development in survival analysis were female sex, older age, higher Body Mass Index, clinically significant parastomal hernia, clinically detectable midline incisional hernia and major postoperative complications ranked as Clavien-Dindo grade III and IV. CONCLUSIONS: Incisional hernia after temporary loop ileostomy is a relevant problem that affects at least one in every ten patients. The previously cited risk factors might favour its development, therefore the use of a prophylactic mesh should be considered in those high-risk patients.

Uva-ursi extract and ibuprofen as alternative treatments for uncomplicated urinary tract infection in women (ATAFUTI): a factorial randomized trial.

OBJECTIVES: The aim was to investigate if offering symptomatic therapy (Uva-ursi or ibuprofen) alongside a delayed prescription would relieve symptoms and reduce the consumption of antibiotics for adult women presenting with acute uncomplicated urinary tract infection (UTI). METHODS: A 2 × 2 factorial placebo controlled randomized trial in primary care. The participants were 382 women aged 18-70 years with symptoms of dysuria, urgency, or frequency of urination and suspected by a clinician to have a lower UTI. The interventions were Uva-ursi extract and/or ibuprofen advice. All women were provided with a delayed or 'back-up' prescription for antibiotics. Missing data were imputed using multiple imputation methods (ISRCTN registry: ISRCTN43397016). RESULTS: An ITT analysis of mean score for frequency symptoms assessed on Days 2-4 found no evidence of a difference between Uva-ursi vs. placebo -0.06 (95% CI -0.33 to 0.21; p 0.661), nor ibuprofen vs. no ibuprofen advice -0.01 (95% CI -0.27 to 0.26; p 0.951). There was no evidence of a reduction in antibiotic consumption with Uva-ursi (39.9% vs. placebo 47.4%; logistic regression odds ratio (OR) 0.59 (95% CI 0.22-1.58; p 0.293) but there was a significant reduction for ibuprofen advice (34.9% vs. no advice 51.0%; OR 0.27 (95% CI 0.10 to 0.72; p 0.009). There were no safety concerns and no episodes of upper tract infection were recorded. CONCLUSIONS: We found no evidence of an effect of either intervention on the severity of frequency symptoms. There is evidence that advice to take ibuprofen will reduce antibiotic consumption without increasing complications. For every seven women given this advice, one less will use antibiotics.

Portal vein arterialization in liver transplantation: an option to restore arterial flow: a case report.

We report a 66-year-old woman who underwent emergency orthotopic liver transplantation due to acute liver failure. The donor's liver graft displayed extensive arteriosclerosis, involving the celiac trunk and hepatic artery. Arterial revascularization of the graft could not be achieved, requiring an arterioportal shunt between the gastroduodenal artery and the portal vein of the recipient. During the early postoperative period, the patient's clinical condition and liver function tests improved rapidly; the patient was discharged on postoperative day 30. Two months later, she developed acute cholangitis. Ischemic-type stenosis of the intrahepatic biliary tree was present, so successful elective retransplantation was undertaken at the ninth postoperative month. In our experience, portal vein arterialization may be useful as a bridging therapy in extreme situations.

Massive low gastrointestinal bleeding due to a Dieulafoy rectal lesion.

Massive low gastrointestinal bleedings are often difficult diagnostically and in terms of management. Sometimes, it is not possible to identify the bleeding point after performing diverse diagnostic tests and the patient undergoes a blind subtotal colectomy. With rectal bleeding, this form of surgery is completely useless, as it will not solve the cause of the haemorrhage. The Dieulafoy lesion has been widely described in the stomach, but in the rectum is a very rare entity that can cause massive lower gastrointestinal bleeding. In the literature, there are only 25 described cases of rectal Dieulafoy lesion.

Posthemipelvectomy hernia.

We report the case of a white male who underwent a classic hemipelvectomy due to a femur fibrosarcoma with inguinal metastases, which 33 years later, developed into a posthemipelvectomy hernia in the amputation stump that impaired the use of his Canadian prosthesis. The hernia was repaired with a polypropylene mesh in a subaponeurotic position. A seroma was drained in the postoperative and it was only 2 months after the operation that he could use his prosthesis with any difficulty. A year after the operation, the hernia had not recurred. Only seven similar cases have been published, and there are only four cases with details of their correction, two with a mesh as was our case, and the rest with a primary suture of the aponeurotic borders. A brief review of the bibliography is given on this subject.

Characterization of the glycosylation profiles of Alzheimer's beta -secretase protein Asp-2 expressed in a variety of cell lines.

Amyloid 39-42 beta -peptides are the main components of amyloid plaques found in the brain of Alzheimer's disease patients. Amyloid 39-42 beta-peptide is formed from amyloid precursor protein by the sequential action of beta- and gamma-secretases. Asp-2 is a transmembrane aspartic protease expressed in the brain, shown to have beta-secretase activity. Mature Asp-2 has four N-glycosylation sites. In this report we have characterized the carbohydrate structures in this glycoprotein expressed in three different cell lines, namely Chinese hamster ovary, CV-1 origin of SV40, and baculovirus-infected SF9 cells. Biantennary and triantennary oligosaccharides of the "complex" type were released from glycoprotein expressed in the mammalian cells, whereas mannose-rich glycans were identified from glycoprotein synthesized in the baculovirus-infected cells. Site-directed mutagenesis of the asparagine residues at amino acid positions 153, 172, 223, and 354 demonstrate that the protease activity of Asp-2 is dependent on its glycosylation.

A G protein-coupled receptor for UDP-glucose.

Uridine 5'-diphosphoglucose (UDP-glucose) has a well established biochemical role as a glycosyl donor in the enzymatic biosynthesis of carbohydrates. It is less well known that UDP-glucose may possess pharmacological activity, suggesting that a receptor for this molecule may exist. Here, we show that UDP-glucose, and some closely related molecules, potently activate the orphan G protein-coupled receptor KIAA0001 heterologously expressed in yeast or mammalian cells. Nucleotides known to activate P2Y receptors were inactive, indicating the distinctly novel pharmacology of this receptor. The receptor is expressed in a wide variety of human tissues, including many regions of the brain. These data suggest that some sugar-nucleotides may serve important physiological roles as extracellular signaling molecules in addition to their familiar role in intermediary metabolism.

Human urotensin-II is a potent vasoconstrictor and agonist for the orphan receptor GPR14.

Urotensin-II (U-II) is a vasoactive 'somatostatin-like' cyclic peptide which was originally isolated from fish spinal cords, and which has recently been cloned from man. Here we describe the identification of an orphan human G-protein-coupled receptor homologous to rat GPR14 and expressed predominantly in cardiovascular tissue, which functions as a U-II receptor. Goby and human U-II bind to recombinant human GPR14 with high affinity, and the binding is functionally coupled to calcium mobilization. Human U-II is found within both vascular and cardiac tissue (including coronary atheroma) and effectively constricts isolated arteries from non-human primates. The potency of vasoconstriction of U-II is an order of magnitude greater than that of endothelin-1, making human U-II the most potent mammalian vasoconstrictor identified so far. In vivo, human U-II markedly increases total peripheral resistance in anaesthetized non-human primates, a response associated with profound cardiac contractile dysfunction. Furthermore, as U-II immunoreactivity is also found within central nervous system and endocrine tissues, it may have additional activities.

Identification and cloning of a connective tissue growth factor-like cDNA from human osteoblasts encoding a novel regulator of osteoblast functions.

We have identified and cloned a novel connective tissue growth factor-like (CTGF-L) cDNA from primary human osteoblast cells encoding a 250-amino acid single chain polypeptide. Murine CTGF-L cDNA, encoding a polypeptide of 251 amino acids, was obtained from a murine lung cDNA library. CTGF-L protein bears significant identity ( approximately 60%) to the CCN (CTGF, Cef10/Cyr61, Nov) family of proteins. CTGF-L is composed of three distinct domains, an insulin-like growth factor binding domain, a von Willebrand Factor type C motif, and a thrombospondin type I repeat. However, unlike CTGF, CTGF-L lacks the C-terminal domain implicated in dimerization and heparin binding. CTGF-L mRNA ( approximately 1.3 kilobases) is expressed in primary human osteoblasts, fibroblasts, ovary, testes, and heart, and a approximately 26-kDa protein is secreted from primary human osteoblasts and fibroblasts. In situ hybridization indicates high expression in osteoblasts forming bone, discrete alkaline phosphatase positive bone marrow cells, and chondrocytes. Specific binding of 125I-labeled insulin-like growth factors to CTGF-L was demonstrated by ligand Western blotting and cross-linking experiments. Recombinant human CTGF-L promotes the adhesion of osteoblast cells and inhibits the binding of fibrinogen to integrin receptors. In addition, recombinant human CTGF-L inhibits osteocalcin production in rat osteoblast-like Ros 17/2.8 cells. Taken together, these results suggest that CTGF-L may play an important role in modulating bone turnover.

Production of monoclonal antibodies in COS and CHO cells.

Recent advances in the generation of genetically engineered monoclonal antibodies have enhanced the importance of COS cells as expression systems for rapidly producing sufficient quantities of these proteins for preliminary biochemical and biophysical analysis. In order to meet the demand for clinical supplies, a gradual increase has occurred in the usage of dihydrofolate reductase negative (DHFR-) Chinese hamster ovary (CHO) cells for large-scale antibody production. Using a variety of mammalian expression vectors and selection/amplification protocols, CHO cell lines capable of producing monoclonal antibodies at levels exceeding 1 gl-1 can now be obtained in an almost routine fashion. For the applications of monoclonal antibodies to expand into additional therapeutic areas, however, a 5-10-fold increase over current highest expression levels may still need to be achieved.

Cloning, expression and functional characterization of type 1 and type 2 steroid 5 alpha-reductases from Cynomolgus monkey: comparisons with human and rat isoenzymes.

The Cynomolgus monkey may provide an alternative pharmacological model in which to evaluate the efficacy of novel inhibitors of the two known human steroid 5 alpha-reductase (SR) isoenzymes. To evaluate the suitability of this species at the level of the molecular targets, a Cynomolgus monkey prostate cDNA library was prepared and screened using human SR type 1 and 2 cDNAs as hybridization probes. Two distinct cDNA sequences were isolated encoding the monkey type 1 and 2 SR isoenzymes. These sequences share 93 and 95% amino acid sequence identity with their human enzyme counterparts, respectively. Difference in monkey type 1 SR, however, was found within the contiguous four amino acids corresponding to the regions in the human and rat sequences that have been proposed previously to influence steroid and inhibitor affinities. Subsequently, both monkey cDNAs were individually expressed in a mammalian cell (CHO) line. Enzyme activities of both monkey SRs were localized to the membrane fractions of CHO cell extracts. Like the human and rat enzymes, the monkey type 1 and type 2 SRs were most active at neutral and low pH, respectively. The results of inhibition studies with over 30 known SR inhibitors, including epristeride, 4MA, and finasteride, indicate that the monkey SR isoenzymes are functionally more similar to the human than the rat homologues. The results from initial velocity and inhibition studies as functions of pH with the human and monkey type 2 SRs also compare favorably. These results, together, suggest that the monkey SR isoenzymes are structurally and functionally comparable on a molecular level to their respective human counterparts, supporting the relevance and use of the Cynomolgus monkey as a pharmacological model for in vivo evaluation of SR inhibitors.

The soluble form of E-selectin is an asymmetric monomer. Expression, purification, and characterization of the recombinant protein.

The gene coding for a soluble form of human E-selectin (sE-selectin) has been expressed in Chinese hamster ovary (CHO) cells. Cells seeded into a hollow fiber reactor secreted protein at a level of 160 mg/liter. The protein was purified to > 95% pure and low endotoxin (< 2 ng/mg), using physiological pH and buffers. The amino acid composition and N-terminal sequence were as predicted from the cDNA sequence. HL-60 cells bound to sE-selectin-coated plates in a dose-dependent manner, and this binding could be blocked up to 100% by pretreatment of HL60 cells with sE-selectin. The concentration of sE-selectin required for 50% inhibition was 1 microM. This value puts an upper limit for the affinity of E-selectin for its natural receptor. sE-selectin also inhibited inflammatory migration of neutrophils in a selective fashion. Purified sE-selectin exhibited a broad band of M(r) approximately 75,000 on nonreducing SDS-PAGE. sE-selectin eluted with M(r) approximately 310,000 from size exclusion chromatography at physiological pH and buffers, suggesting an oligomeric state. Matrix-assisted laser-desorption MS gave a molecular weight of 80,000, while the minimum monomer molecular weight from the gene sequence should be 58,571, demonstrating that the monomeric molecule thus expressed had 27% carbohydrate. Equilibrium analytical ultracentrifugation gave an average solution molecular weight of 81,600 (+/- 4,500). Velocity ultracentrifugation gave a sedimentation coefficient of 4.3 S and, from this, an apparent axial ratio of 10.5:1, assuming a prolate ellipsoid of revolution. An analysis of the NMR NOESY spectra of sE-selectin, sialyl-Lewis X, and sE-selectin with sialyl-Lewis X demonstrates that the recombinant protein binds sialyl-Lewis X productively. Hence, in solution, sE-selectin is a functional elongated monomer.

Human AT1 receptor is a single copy gene: characterization in a stable cell line.

To address conflicting reports concerning the number of angiotensin II (AII) receptor type 1 (AT1) coding loci in vertebrates, Southern blot analysis was used to determine the genomic representation of AT1 receptor genes in animals comprising a divergent evolutionary spectrum. The data demonstrate that the AT1 receptor gene is present as a single genomic copy in a broad spectrum of animals including human, monkey, dog, cow, rabbit, and chicken. In contrast, members of the rodent taxonomic order contain two genes in their genomes. These two genes may have arisen in rodents as a consequence of a gene duplication event that occurred during evolution following the branching of rodents from the mammalian phylogenetic tree. In order to investigate the properties of the human AT1 receptor in a pure cell system, the recombinant human AT1 receptor was stably expressed in mouse L cells. An isolated cell line, designated LhAT1-D6, was found to express abundant levels of recombinant receptor [430 +/- 15 fmol/mg] exhibiting high affinity [KD = 0.15 +/- 0.02 nM] for [125I][SAR1, Ile8] angiotensin II (SIA). The pharmacological profile of ligands competing for [125I] SIA binding to the expressed receptor was in accordance with that of the natural receptor. Radioligand binding of the expressed receptor was decreased in the presence of the non-hydrolyzable analog of GTP, guanosine 5'-(gamma-thio) triphosphate [GTP gamma S]. Angiotensin II evoked a rapid efflux of 45Ca2+ from LhAT1-D6 cells that was blocked by AT1 receptor specific antagonists. In addition, AII inhibited forskolin-stimulated cAMP accumulation in these cells which was blocked by the AT-1 antagonist. Thus, the LhAT1-D6 cell line provides a powerful tool to explore the human AT1 receptor regulation.

DHFR coamplification of t-PA in DHFR+ bovine endothelial cells: in vitro characterization of the purified serine protease.

High-level expression of human tissue-type plasminogen activator was accomplished in endothelial cells by a novel approach to dihydrofolate reductase (DHFR) coamplification in DHFR+ cells. A tripartite mammalian expression vector coding for DHFR, neomycin phosphotransferase, and the t-PA gene was introduced into bovine endothelial cells by transfection and selection for G418 resistance. Upon methotrexate selection of these transformants, we obtained endothelial cells that had amplified the plasmid-encoded DHFR and t-PA genes. As a result, cell lines were isolated that efficiently produced t-PA (greater than 4 pg/cell.day). This t-PA was purified and compared with recombinant t-PA produced in Chinese hamster ovary cells. These two t-PA samples differed in carbohydrate composition, and amounts of 530 and 527 amino acid forms but had similar in vitro activity.

Use of two different deoxyribonucleic acid probes to detect Y chromosome deoxyribonucleic acid in subjects with normal and altered Y chromosomes.

Four experiments evaluated the sensitivity and specificity of molecular techniques to detect human Y chromosome deoxyribonucleic acid. In experiment I, electrophoretic separation of normal male deoxyribonucleic acid fragments after digestion with endonuclease Hae III revealed two male-specific bands of 3.4 and 2.1 kilobase (kb). These bands were not visible if the fraction of male deoxyribonucleic acid in mixed samples was less than 0.3. In experiment II, by means of a repetitive copy Y deoxyribonucleic acid probe (pS4) mapped to Yq12, a male-specific 2.3 kb band was detectable in mixtures of 2.5 ng of male deoxyribonucleic acid and 997.5 ng of 45,X female deoxyribonucleic acid. In experiment III, hybridization with the pS4 probe was performed on the deoxyribonucleic acid of 20 subjects with a normal or a variant Y chromosome. In experiment IV, deoxyribonucleic acid from the same subjects was hybridized to a single copy probe (4B-2) mapped to the Yq11 region. Deoxyribonucleic acid from category A subjects (n = 8) with cytologically normal Y chromosomes hybridized to both deoxyribonucleic acid probes. Deoxyribonucleic acid from category B subjects (n = 2), including a variant Y chromosome that was negative for Q-banding but positive for C-bands, hybridized with the distal pS4 and proximal 4B-2 probes. Deoxyribonucleic acid from category C subjects (n = 10) with variant Y chromosomes uniformly negative for Q- and C-bands, did not hybridize with the pS4 probe. Deoxyribonucleic acid from three of the 10 category C subjects did hybridize to the more proximal sequence-detecting 4B-2 probe. Deoxyribonucleic acid from the remaining seven subjects in category C did not hybridize with either of the deoxyribonucleic acid probes.