Association of preoperative anaemia with postoperative morbidity and mortality: an observational cohort study in low-, middle-, and high-income countries.

BACKGROUND: Anaemia is associated with poor postoperative outcomes, but few studies have described the impact of preoperative anaemia in low- and middle- (LMICs), and high-income countries (HICs). METHODS: This was a planned analysis of data collected during an international 7 day cohort study of adults undergoing elective inpatient surgery. The primary outcome was in-hospital death, and the secondary outcomes were in-hospital complications. Anaemia was defined as haemoglobin <12 g dl-1 for females and <13 g dl-1 for males. Hierarchical three-level mixed-effect logistic regression models were constructed to examine the associations between preoperative anaemia and outcomes. RESULTS: We included 38 770 patients from 474 hospitals in 27 countries of whom 11 675 (30.1%) were anaemic. Of these, 6886 (17.8%) patients suffered a complication and 198 (0.5%) died. Patients from LMICs were younger with lower ASA physical status scores, but a similar prevalence of anaemia [LMIC: 5072 (32.5%) of 15 585 vs HIC: 6603 (28.5%) of 23 185]. Patients with moderate [odds ratio (OR): 2.70; 95% confidence interval (CI): 1.88-3.87] and severe anaemia (OR: 4.09; 95% CI: 1.90-8.81) were at an increased risk of death in both HIC and LMICs. Complication rates increased with the severity of anaemia. Compared with patients in LMICs, those in HICs experienced fewer complications after an interaction term analysis [LMIC (OR: 0.92; 95% CI: 0.87-0.97) vs HIC (OR: 0.86; 95% CI: 0.84-0.87); P<0.01]. CONCLUSIONS: One-third of patients undergoing elective surgery are anaemic. These patients have an increased risk of complications and death. The prevalence of anaemia is similar amongst patients in LMICs despite their younger age and lower risk profile. CLINICAL TRIAL REGISTRATION: ISRCTN51817007.

Comparison of the effects of lactated Ringer solution with and without hydroxyethyl starch fluid resuscitation on gut edema during severe splanchnic ischemia.

The type of fluid used during resuscitation may have an important impact on tissue edema. We evaluated the impact of two different regimens of fluid resuscitation on hemodynamics and on lung and intestinal edema during splanchnic hypoperfusion in rabbits. The study included 16 female New Zealand rabbits (2.9 to 3.3 kg body weight, aged 8 to 12 months) with splanchnic ischemia induced by ligation of the superior mesenteric artery. The animals were randomized into two experimental groups: group I (N = 9) received 12 mL x kg-1 x h-1 lactated Ringer solution and 20 mL/kg 6% hydroxyethyl starch solution; group II (N = 7) received 36 mL x kg-1 x h-1 lactated Ringer solution and 20 mL/kg 0.9% saline. A segment from the ileum was isolated to be perfused. A tonometric catheter was placed in a second gut segment. Superior mesenteric artery (Q SMA) and aortic (Qaorta) flows were measured using ultrasonic flow probes. After 4 h of fluid resuscitation, tissue specimens were immediately removed for estimations of gut and lung edema. There were no differences in global and regional perfusion variables, lung wet-to-dry weight ratios and oxygenation indices between groups. Gut wet-to-dry weight ratio was significantly lower in the crystalloid/colloid-treated group (4.9 +/- 1.5) than in the crystalloid-treated group (7.3 +/- 2.4) (P < 0.05). In this model of intestinal ischemia, fluid resuscitation with crystalloids caused more gut edema than a combination of crystalloids and colloids.

Comparison of the effects of lactated Ringer solution with and without hydroxyethyl starch fluid resuscitation on gut edema during severe splanchnic ischemia

The type of fluid used during resuscitation may have an important impact on tissue edema. We evaluated the impact of two different regimens of fluid resuscitation on hemodynamics and on lung and intestinal edema during splanchnic hypoperfusion in rabbits. The study included 16 female New Zealand rabbits (2.9 to 3.3 kg body weight, aged 8 to 12 months) with splanchnic ischemia induced by ligation of the superior mesenteric artery. The animals were randomized into two experimental groups: group I (N = 9) received 12 mL·kg-1·h-1 lactated Ringer solution and 20 mL/kg 6 percent hydroxyethyl starch solution; group II (N = 7) received 36 mL·kg-1·h-1 lactated Ringer solution and 20 mL/kg 0.9 percent saline. A segment from the ileum was isolated to be perfused. A tonometric catheter was placed in a second gut segment. Superior mesenteric artery (Q SMA) and aortic (Qaorta) flows were measured using ultrasonic flow probes. After 4 h of fluid resuscitation, tissue specimens were immediately removed for estimations of gut and lung edema. There were no differences in global and regional perfusion variables, lung wet-to-dry weight ratios and oxygenation indices between groups. Gut wet-to-dry weight ratio was significantly lower in the crystalloid/colloid-treated group (4.9 ± 1.5) than in the crystalloid-treated group (7.3 ± 2.4) (P < 0.05). In this model of intestinal ischemia, fluid resuscitation with crystalloids caused more gut edema than a combination of crystalloids and colloids.

RF tumour ablation: computer simulation and mathematical modelling of the effects of electrical and thermal conductivity.

This study determined the effects of thermal conductivity on RF ablation tissue heating using mathematical modelling and computer simulations of RF heating coupled to thermal transport. Computer simulation of the Bio-Heat equation coupled with temperature-dependent solutions for RF electric fields (ETherm) was used to generate temperature profiles 2 cm away from a 3 cm internally-cooled electrode. Multiple conditions of clinically relevant electrical conductivities (0.07-12 S m-1) and 'tumour' radius (5-30 mm) at a given background electrical conductivity (0.12 S m-1) were studied. Temperature response surfaces were plotted for six thermal conductivities, ranging from 0.3-2 W m-1 degrees C (the range of anticipated clinical and experimental systems). A temperature response surface was obtained for each thermal conductivity at 25 electrical conductivities and 17 radii (n=425 temperature data points). The simulated temperature response was fit to a mathematical model derived from prior phantom data. This mathematical model is of the form (T=a+bRc exp(dR) s(f) exp(g)(s)) for RF generator-energy dependent situations and (T=h+k exp(mR)+n?exp(p)(s)) for RF generator-current limited situations, where T is the temperature (degrees C) 2 cm from the electrode and a, b, c, d, f, g, h, k, m, n and p are fitting parameters. For each of the thermal conductivity temperature profiles generated, the mathematical model fit the response surface to an r2 of 0.97-0.99. Parameters a, b, c, d, f, k and m were highly correlated to thermal conductivity (r2=0.96-0.99). The monotonic progression of fitting parameters permitted their mathematical expression using simple functions. Additionally, the effect of thermal conductivity simplified the above equation to the extent that g, h, n and p were found to be invariant. Thus, representation of the temperature response surface could be accurately expressed as a function of electrical conductivity, radius and thermal conductivity. As a result, the non-linear temperature response of RF induced heating can be adequately expressed mathematically as a function of electrical conductivity, radius and thermal conductivity. Hence, thermal conductivity accounts for some of the previously unexplained variance. Furthermore, the addition of this variable into the mathematical model substantially simplifies the equations and, as such, it is expected that this will permit improved prediction of RF ablation induced temperatures in clinical practice.

Effects of maximizing oxygen delivery on morbidity and mortality in high-risk surgical patients.

OBJECTIVE: To evaluate the effects of maximizing the oxygen delivery on morbidity and mortality in patients >60 yrs of age and/or with chronic diseases of vital organs who underwent major elective surgery. DESIGN: Prospective, randomized, controlled trial. SETTING: A 24-bed general intensive care unit of a teaching hospital. PATIENTS: Thirty-seven high-risk patients who underwent major surgery. INTERVENTIONS: The hemodynamic and oxygen transport variables and outcomes in 18 patients (control group) treated to maintain normal values of oxygen delivery were compared with 19 patients (protocol group) treated to maintain "supranormal" values. Therapy in both groups consisted of volume expansion and, when necessary, dobutamine to reach target values, during the surgery and 24 hrs postoperatively. MEASUREMENTS AND MAIN RESULTS: We interrupted the study because of a significant difference in the 60-day mortality rate. The mortality rate in the control group was significantly higher when compared with the protocol group (9/18 [50%] vs. 3/19 [15.7%], p < .05). The prevalence of clinical and infectious complications was higher in the control group than in the protocol group (67% and 31% respectively; relative risk, 0.47; 95% confidence interval, 0.226-0.991; p < .05) and there was a trend toward more severe organ dysfunction in nonachievers patients (17/24 [71%] vs. 6/13 [46%], relative risk, 0.65; 95% confidence interval, 0.343-1.237; NS). CONCLUSION: Older patients with existing cardiorespiratory illness undergoing major surgery have a reduced morbidity and mortality when dobutamine is used to maximize oxygen transport.

Targeting TBP to a non-TATA box cis-regulatory element: a TBP-containing complex activates transcription from snRNA promoters through the PSE.

In the human small nuclear RNA (snRNA) promoters, the presence of a TATA box recognized by the TATA box-binding protein (TBP) determines the selection of RNA polymerase III over RNA polymerase II. The RNA polymerase II snRNA promoters are, therefore, good candidates for TBP-independent promoters. We show here, however, that TBP activates transcription from RNA polymerase II snRNA promoters through a non-TATA box element, the snRNA proximal sequence element (PSE), as part of a new snRNA-activating protein complex (SNAPc). In contrast to the previously identified TBP-containing complexes SL1, TFIID, and TFIIIB, which appear dedicated to transcription by a single RNA polymerase, SNAPc is also essential for RNA polymerase III transcription from the U6 snRNA promoter. The U6 initiation complex appears to contain two forms of TBP, one bound to the TATA box and one bound to the PSE as a part of SNAPc, suggesting that multiple TBP molecules can have different functions within a single promoter.

A TBP complex essential for transcription from TATA-less but not TATA-containing RNA polymerase III promoters is part of the TFIIIB fraction.

The TATA box-binding protein TBP directs transcription by all three eukaryotic RNA polymerases. In mammalian cells, TBP is found in at least three different complexes: SL1, D-TFIID, and B-TFIID. While SL1 and D-TFIID are involved in RNA polymerase I and II transcription, respectively, no unique function has been assigned to the B-TFIID complex. Here we show that the TFIIIB fraction required for RNA polymerase III transcription contains two separable components, one of which is a TBP-containing complex that may correspond to B-TFIID. For transcription of TATA-less RNA polymerase III genes such as the VAI, 5S, and 7SL genes, this complex cannot be replaced by either TBP alone or the D-TFIID complex. In contrast, TBP alone is active for basal transcription from the TATA-containing U6 promoter. This indicates different requirements for recruiting TBP to TATA-less and TATA-containing RNA polymerase III promoters.

The cloned RNA polymerase II transcription factor IID selects RNA polymerase III to transcribe the human U6 gene in vitro.

Although the human U2 and U6 snRNA genes are transcribed by different RNA polymerases (i.e., RNA polymerases II and III, respectively), their promoters are very similar in structure. Both contain a proximal sequence element (PSE) and an octamer motif-containing enhancer, and these elements are interchangeable between the two promoters. The RNA polymerase III specificity of the U6 promoter is conferred by a single A/T-rich element located around position -25. Mutation of the A/T-rich region converts the U6 promoter into an RNA polymerase II promoter, whereas insertion of the A/T-rich region into the U2 promoter converts that promoter into an RNA polymerase III promoter. We show that this A/T-rich element can be replaced by a number of TATA boxes derived from mRNA promoters transcribed by RNA polymerase II with little effect on RNA polymerase III transcription. Furthermore, the cloned RNA polymerase II transcription factor TFIID both binds to the U6 A/T-rich region and directs accurate RNA polymerase III transcription in vitro. Mutations in the U6 A/T-rich region that convert the U6 promoter into an RNA polymerase II promoter also abolish TFIID binding. Together, these observations suggest that in the human snRNA promoters, unlike in mRNA promoters, binding of TFIID directs the assembly of RNA polymerase III transcription complexes, whereas the lack of TFIID binding results in the assembly of RNA polymerase II snRNA transcription complexes.

cis-acting elements required for RNA polymerase II and III transcription in the human U2 and U6 snRNA promoters.

Although the human U2 and U6 snRNA genes are transcribed by RNA polymerases II and III respectively, their promoters are remarkably similar in structure. Both promoters contain a proximal element and an enhancer region with an octamer motif. The U6 promoter contains in addition an A/T rich region that defines it as an RNA polymerase III promoter. We have examined in further detail the contributions of sequences in the human U2 and U6 promoter regions to transcription by RNA polymerase II and III. We find that although the sequences surrounding the U2 cap site favor RNA polymerase II transcription, their presence cannot suppress a shift to RNA polymerase III specificity upon insertion of the U6 A/T box. In the U6 promoter, the 3' part of the proximal element homology is essential for efficient transcription and is also involved in localizing the start site of transcription. A region downstream of the proximal element homology is required for RNA polymerase II (but not for RNA polymerase III) transcription, both in the U2 promoter and in the U6 promoter. This element may be recognized by an RNA polymerase II transcription factor or by RNA polymerase II itself. The presence of this element in the U6 promoter raises the possibility that the human U6 gene is, under certain circumstances, transcribed by RNA polymerase II.

U3 snRNPs and nucleolar development during oocyte maturation, fertilization and early embryogenesis in the mouse: U3 snRNA and snRNPs are not regulated coordinate with other snRNAs and snRNPs.

U3 small nuclear ribonucleic acids (snRNA) and U3 small nuclear ribonucleoprotein (snRNP), which are thought to be responsible for ribosomal RNA processing, are quantitated and localized during oocyte maturation, fertilization, and early embryogenesis in the mouse. On the basis of Northern blot and nuclease protection experiments, it is estimated that there are about 5 x 10(4) U3 snRNA molecules in an ovulated oocyte and in a two-cell embryo. This number then increases roughly 50-fold to 2.7 x 10(6) molecules per embryo by the blastocyst stage. At all stages of development U3 snRNP antigens colocalize with nucleoli, as defined by differential interference contrast microscopy and an antibody to a nucleolar epitope. The synthesis and distribution of U3 snRNA and U3 snRNP follow a pattern independent from other major U snRNPs and snRNAs.

A 7 bp mutation converts a human RNA polymerase II snRNA promoter into an RNA polymerase III promoter.

The human U2 snRNA promoter directs the formation of a specialized RNA polymerase II transcription complex that recognizes the snRNA gene 3' box as a signal for RNA 3' end formation. In contrast, the human U6 promoter is recognized by RNA polymerase III and transcription terminates in a run of Ts. We show that transcription from the U6 promoter is dependent on a sequence similar to the U2 proximal element and on an AT-rich element centered around position -27. Mutation of the AT-rich element induces RNA polymerase II transcription from the U6 promoter, whereas insertion of this element within the U2 promoter converts it into a predominantly RNA polymerase III promoter. The site of transcription termination always correlates with the nature of the transcribing polymerase: the 3' box with RNA polymerase II and a run of Ts with RNA polymerase III. Thus, a single element determines the RNA polymerase specificity of snRNA promoters and hence the site of transcription termination.

Localization and expression of U1 RNA in early mouse embryo development.

We have studied the accumulation and localization of U1 RNA during mouse embryo development by in situ hybridization with a U1 RNA probe and immunofluorescence microscopy using a mouse monoclonal antibody to U1 snRNP. There is a substantial amount of U1 RNA present in the oocyte that is present in both the germinal vesicle and the cytoplasm although the concentration is higher in the nuclear compartment. Following the germinal vesicle breakdown that accompanies ovulation and meiotic maturation, the U1 RNA is uniformly distributed throughout the unfertilized oocyte. In the fertilized egg, the silver grain density from in situ hybridization is higher over pronuclei and this enrichment is maintained at the two-cell and later stages. Similar results were obtained for the distribution of the U1 snRNP as assayed by immunofluorescence microscopy: U1 RNA is predominantly localized in all nuclei except polar body nuclei. The U1 RNA in the oocyte and two-cell embryo is predominantly (greater than 85%) U1a RNA. By the eight-cell stage there is a two to three-fold increase in the amount of total U1 RNA and the proportion of U1b RNA has increased to about 40%. The amount of U1 RNA continues to increase through the blastocyst stage and the proportion of the U1b RNA increases to 60%.

Synthesis of U1 RNA in isolated mouse cell nuclei: initiation and 3'-end formation.

The transcription of U1 RNA genes was studied in isolated nuclei from mouse myeloma cells. Using a cloned U1b gene as a probe, we showed that isolated nuclei synthesize both U1b and U1a RNA. The U1 RNAs were initiated in vitro, as measured by incorporation of adenosine 5'-O-(2-thiotriphosphate) into U1 RNA. There was transcription of the 3'-flanking region but no transcription of regions directly 5' to the U1 genes. In addition to U1 RNAs of the correct length which were released from the nuclei, there were larger RNAs, presumably resulting from transcription into the 3'-flanking region, which were retained in the nuclei. Chase experiments showed that these longer transcripts were not precursors to mature U1 RNA, a finding consistent with the idea that 3'-end formation is coincident with transcription. During the chase, there was maturation of the 3' ends of U1a and U1b RNAs from slightly longer precursors. In addition to accurate transcription of U1 RNA, there was also synthesis of U2 and U3 RNA. All three of these RNAs were transcribed by RNA polymerase II, as measured by their sensitivity to alpha-amanitin.

Expression of a mouse U1b gene in mouse L cells.

A 6.9-kb DNA fragment containing two mouse U1b genes was introduced by cotransfection with the Herpes simplex virus (HSV) thymidine kinase (TK) gene into tk- mouse L cells. The parent Ltk- cells produce primarily U1a RNA and only small amounts of U1b RNA. However, after introduction of exogenous U1b genes by cotransfection these cells express large amounts of U1b RNA. Subcloned DNA fragments containing a single U1b gene and varying amounts of DNA flanking the 5' -end of the gene were introduced into Ltk- cells. All of the constructs containing 400 bp or more upstream of the U1b gene were efficiently expressed. By comparison, a subclone containing only 150 bp of DNA flanking the 5' -end of the U1b gene was not efficiently expressed. The U1b transcripts synthesized in transfected cells were identified by hybrid selection and by precipitation of ribonucleoproteins (RNP) containing U1 RNA with anti-RNP antibody.