Primary Structure of a Trypsin Inhibitor (Copaifera langsdorffii Trypsin Inhibitor-1) Obtained from C. langsdorffii Seeds.

In this study, the aim was to determine the complete sequence of the Copaifera langsdorffii trypsin inhibitor (CTI)-1 using 2-dimensional (2D)-PAGE, matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF), and quadrupole time-of-flight (QTOF) spectrometry. Spots A (CTI-1) and F (CTI-2) were submitted to enzymatic digestions with trypsin, SV8, and clostripain. The accurate mass of the peptide obtained from each digest was determined by mass spectrometry (MS) using MALDI-TOF. The most abundant peptides were purified and sequenced in a liquid chromatograph connected to an electrospray ionization-QTOF MS. When the purified trypsin inhibitor was submitted to 2D electrophoresis, different spots were observed, suggesting that the protein is composed of 2 subunits with microheterogeneity. Isoelectric points of 8.0, 8.5, and 9.0 were determined for the 11 kDa subunit and of 4.7, 4.6, and 4.3 for the 9 kDa subunit. The primary structure of CTI-1, determined from the mass of the peptide of the enzymatic digestions and the sequence obtained by MS, indicated 180 shared amino acid residues and a high degree of similarity with other Kunitz (KTI)-type inhibitors. The peptide also contained an Arg residue at the reactive site position. Its 3-dimensional structure revealed that this is because the structural discrepancies do not affect the canonical conformation of the reactive loop of the peptide. Results demonstrate that a detailed investigation of the structural particularities of CTI-1 could provide a better understanding of the mechanism of action of these proteins, as well as clarify its biologic function in the seeds. CTI-1 belongs to the KTI family and is composed of 2 polypeptide chains and only 1 disulfide bridge.

Microvascular autonomic dysfunction may justify false-positive stress myocardial perfusion imaging in patients with liver cirrhosis undergoing liver transplantation.

BACKGROUND: Up to 15% of liver transplant candidates have asymptomatic coronary artery diseases, which increase the risk of cardiac complications during and after transplantation. The aim of this study was to prospectively investigate the usefulness of an integrated cardiological approach in cirrhotic patients undergoing liver transplantation. METHODS: Twenty-four consecutive patients undergoing evaluation for liver transplantation were studied by assessing risk factors for coronary artery diseases, electrocardiogram with QTc interval determination, chest X-ray, echocardiography, 24-hour Holter monitor, myocardial perfusion scintigraphy (99mTc)MIBI-GSPECT at rest and after dipyridamole infusion. Cardiac (123)I-metaiodobenzylguanidine (MIBG) scan and coronarography were performed in patients with myocardial perfusion defects. Twenty three of 24 patients underwent successful liver transplantation; one patient died on the waiting list. RESULTS: Before liver transplantation, 29% of patients were diabetic and 41% were smokers. Eleven of 24 patients had a prolonged QTc interval, and 3/24 had positive myocardioscintigraphy after dipyridamole infusion: in two coronarography was negative, while the (123)I-MIBG washout was altered. No cardiac events were recorded during the short-and long-term follow-up after surgery. CONCLUSIONS: Predictive value of positive cardiac (99mTc)MIBI-GSPECT in patients with liver cirrhosis is low, and this may be due to alterations of cardiac microvascular tone as showed by cardiac (123)I-MIBG scan.

Activity of GH/IGF-I axis in trauma and septic patients during artificial nutrition: different behavior patterns?

The aim of this study was to compare several parameters of GH/IGF-I axis activity in septic and trauma patients during Intensive Care Unit (ICU) stay. To this goal, 13 patients with sepsis (SEP) and 16 with trauma (TRA) were studied. Thirty-three adult subjects (AS) were studied as controls. Serum IGF-I and -II, IGFBP-1, -2 and -3, GH and GHBP levels were studied on day 1, 3, 5 and 7 after ICU admission, during comparable artificial nutrition in SEP and TRA and basally in AS. In 5 patients with SEP and 6 with TRA, the GH response to GHRH was evaluated on day 3. On ICU day 1, IGF-I and -II and IGFBP-3 in SEP were lower (p<0.05) than in TRA which, in turn, were lower (p<0.01) than in AS. IGF-I increased (p<0.05) both in SEP and TRA, but, on ICU day 7, those in SEP persisted lower than in TRA, which became similar to those in AS. IGF-II levels increased (p<0.05) in SEP only, persisting lower (p<0.05) than in TRA. On ICU day 1, GH in SEP and TRA were similar and did not vary until day 7, overlapping those in AS. The GH response to GHRH in SEP and TRA was similar and lower (p<0.01) than in AS. These findings indicate that IGF-I activity is impaired more in septic than in trauma patients. Reduced IGF-I activity probably reflects peripheral GH resistance though basal and GHRH-induced GH levels were not increased in these conditions.

Energy expenditure (EE) and substrate utilization (SU) in the perioperative period in orthotopic liver transplantation (OLTX).

12 patients were investigated with IC (Datex, Deltatrac) preoperatively and during the surgical procedure of OLTX, VO2, VCO2 were continuously measured and RQ, EE and SU were calculated considering the different periods of the procedure: preoperative resting EE: (PREE), anaesthesiological procedures (ANEE), liver preparation EE (LPEEE), liver removal EE (LREE), anhepatic phase EE (APEE), reperfusion, EE (RPEE) and end of operation EE (EOEE). EE were expressed as % respect value calculated with H.B. (Harris-Benedict) formula. Data were analyzed with Student T-test and p < 0.01** or < 0.05* PREE is typical in end stage liver disease with low RQ values and increased EE. Energy production depends on lipid utilization since liver gluconeogenesis and glycogen stores are impaired. Anesthesia reduces energy needs and production up to 50% of the preoperative values, reducing VO2 more than VCO2 and therefore an augmented RQ value over 1.0. SU analysis indicates an increased glucose and aminoacid utilization coupled with high nitrogen catabolism that continues in the postoperative period (from 0.08 0.01 gN/kg b.w. to 0.20 0.06 gN/kg b.w.). When the new liver is reperfused, VO2 increases more than VCO2 indicating the risk of reperfusion injury.

[Subarachnoid anesthesia with Sprotte 24-G needle in ambulatory surgery]. / Anestesia subaracnoidea con ago di Sprotte 24-G in chirurgia ambulatoriale.

In order to evaluate the possibility of using a selective subarachnoid anesthesia in ambulatory surgery we studied its feasibility with a Sprotte needle 24 G in 103 patients (range 15-67 years) undergoing a knee joint arthroscopy. The subarachnoid anesthesia was achieved with Sprotte needle 24 G with introducer with patient on lateral decubitus. Hyperbaric bupivacaine 1% (0.05 mg/cm height) was used in order to obtain a selective homolateral metameric anesthesia between L1 and S3. The numbers of attempts were 1.29 (range 1-4). We involved the homolateral determatomeres in 94% of our attempts, adding a weak anesthetic action on contralateral ones. In the remaining 6% of the patients the anesthesia was bilateral and extending in an overlying manner. Anesthesia was adequate for the time needed to perform all the surgical procedures. Nevertheless in 3 patients the tourniquet was painful. We registered systemic hypotension only in 2% of the patients (a reduced systolic pressure value > 30%). No cases of postdural puncture headache were registered. Non postural atypical headache was seem in 3.9% of the patients. Urinary retention necessitating catheterization was noted only in 3 cases. A weak backache was revealed in 8.8%. When interviewed after surgery, 98% of our patients fulfilled the technique. In spite of technical difficulties that the use of the Sprotte needle may present, the selective subarachnoid anesthesia may be a safe alternative to the other anesthetic procedures for ambulatory surgery of the legs.

TSF, RBP and TBPA as nutritional indices in surgical patients with gynecological cancer.

The Authors investigated the usefulness of some biochemical markers of visceral protein synthesis (TSF, TBPA and RBP) in 24 patients affected with gynecological cancer and treated with Total Parenteral Nutrition in the perioperative period. The absence of an improving TSF and TBPA is related to increased morbidity and mortality.

Early outcome prediction in severe head injury: comparison between children and adults.

The prognostic value in 49 children and 56 adults of the following parameters recorded on admission was analyzed: oculocephalic and light reflexes, posturing, Glasgow Coma Scale (GCS), the need for ventilatory support, and the presence of associated injuries. All patients had been in coma for at least 6 h. The presence of intracranial hematoma and the duration of coma were recorded and the relative risk of poor outcome calculated. There was poor outcome in 51% of the children and 61% of the adults. Oculocephalic and light reflexes, posturing, GCS, need for ventilatory support, and duration of coma were significantly related to the outcome in children. Only oculocephalic and light reflexes, and posturing were significantly related to the outcome in adults. Some parameters appeared to have different prognostic value in children and in adults: the simultaneous evaluation of oculocephalic reflex and need for ventilation was the best prognostic guide in children, the light reflex was the best prognostic indicator in adults.