alpha1-Acid glycoprotein production in rat dorsal air pouch in response to inflammatory stimuli, dexamethasone and honey bee venom.

This study shows the rapid and differential production of the 40-43 kDa and the 70-90 kDa alpha1-acid glycoprotein (AGP) fucosylated glycoforms after treatment of the dorsal air pouch with bacterial lipopolysaccharide (LPS), HgCl(2) or Freund's complete adjuvant (FCA). The 40-43 kDa and the 70-90 kDa AGP production is peaked 1-3 h post-LPS treatment. We observed that the responses to LPS and FCA are similar in that both AGP isoforms are induced whereas they differ in that the FCA exhibits a 6 h lag period. The response to HgCl(2,) however, exhibits the specific biphasic induction only of the 40-43 kDa AGP. The serum 40-43 kDa AGP glycoform gradually increases in response to all of the above stimulants and peaks by 24 h post- treatment. The increase of the 70-90 kDa AGP levels in the air pouch occurs in association with the accumulation of polymorphonuclear (PMN) cells while dexamethasone (DEX) increases only the 40-43 kDa AGP production in the absence of PMN accumulation. Macrophage-monocyte lineage cells forming the air pouch lining tissue may potentially be the cells that secrete the 40-43 kDa AGP while polymorphonuclear cells that infiltrate the air pouch secrete the 70-90 kDa AGP. The 40-43 kDa and 70-90 kDa AGP production induced by LPS in the air pouch precedes that of interleukin-1 (IL-1) or interleukin-6 (IL-6) while the 40-43 kDa AGP glycoform potentially increases IL-6 production by air pouch PMN exudate cells. These significant differences suggest a local pro-inflammatory role of AGP. Honeybee venom suppressed arthritis development and exhibited differential local or systemic regulation of AGP in serum vs. air pouch exudate or synovial fluid. This study with the air pouch model of facsimile synovium tissue suggests that local alpha1-acid glycoprotein (AGP) production may contribute to pro-inflammatory and anti-inflammatory activities during the local acute phase response or during chronic inflammatory stress as in arthritis.

Validation of the dorsal air pouch model to predict and examine immunostimulatory responses in the gut.

AIMS: To validate the use of the air pouch system to predict and examine early immune responses induced by the presumptive probiotics Lactobacillus paracasei subsp. paracasei B112, DC205, DC215 and DC412 strains in the gut mucosa. METHODS AND RESULTS: Only the DC412 strain interacted strongly with the cells forming the air pouch lining tissue and induced early innate immune responses such as polymorphonuclear (PMN) cell recruitment, phagocytosis and tumour necrosis factor alpha (TNF-alpha) production that equal the respective responses induced by the probiotic Lactobacillus acidophilus NCFB 1748. The strains exhibiting strong immunoregulatory activity in the air pouch also interacted strongly with the gut-associated lymphoid tissue (GALT). The strain DC412 exerts its effect on the intestine through stimulation of Toll-like receptor (TLR)2/TLR4-mediated signalling events leading to secretion of a certain profile of cytokines in which gamma interferon (IFN-gamma), TNF-alpha, interleukin (IL)-6 and IL-10 are included. The probiotic Lact. acidophilus NCFB 1748 induces the same cytokine profile in addition to IL-12B, and this response is potentially mediated by the synergy of TLR2 and TLR9. CONCLUSION: The strain DC412 possesses the in vitro and in vivo characteristics of a probiotic micro-organism. SIGNIFICANCE AND IMPACT OF THE STUDY: The dorsal mouse or rat air pouch may be used as an alternative and rapid method for the initial discrimination and selection of potential probiotic Lactobacillus strains.

Conditioned narcotic withdrawal in humans.

Subjective and physiological manifestations of the narcotic withdrawal syndrome were produced as a conditioned response. Withdrawal reactions precipitated by the narcotic antagonist naloxone in methadone-dependent volunteers were the unconditioned response. These data support clinical anecdotes of withdrawal symptoms occurring in former addicts when they return to their drug-related environment.

Proteolysis of milk fat globule membrane proteins in preterm milk: a transient phenomenon with a possible biological role?

Milk fat globule membrane (MFGM) proteins constitute a milk fraction currently of great interest, as they appear to significantly contribute to milk protective role. We investigated these proteins in human preterm colostrum and milk. For the former we found a peculiar 2-DE pattern, with a spot concentration at low molecular weight, which mass spectrometry analysis showed to be fragments belonging to some MFGM proteins with a well-known biological and especially immunological role: lactadherin, membrane-associated lactoferrin, butyrophilin, clusterin and heavy-chain immunoglobulin. Since we were able to rule out protease activity after specimen collection, we hypothesize the localization of the proteolytic enzymes in the alveolar cell membranes of the mammary gland. This mechanism is probably under hormonal control and the unexpected advent of preterm delivery would not allow hormonal conditions typical of lactation to occur immediately, causing a delay in enzymatic inhibition. This hypothesis is supported by some of our results, picturing a peculiar transient phenomenon of adaptation of the mammary-gland-membrane proteins after preterm delivery. Further studies will be required to verify whether the presence of protein fragments exerts a specific biological and immuno-defensive role in preterm infants, thus adding evidence to the outstanding biological role and benefits of mother's own milk in feeding preterm infants.

Effects of Holder pasteurization on human milk oligosaccharides.

The benefits of human milk have been confirmed for preterm infants, due to its nutritional aspects and to its biologically active compounds. Oligosaccharides play an emerging leading role among these compounds. Mother's milk can sometimes be lacking for preterm infants; pasteurized donor milk represents therefore an important alternative. The aim of this study is to evaluate the effects of Holder pasteurization on the concentration and pattern of oligosaccharides in preterm human milk. Our results indicate that pasteurization does not affect the concentration or pattern of analyzed oligosaccharides.

Immunoglobulin-A profile in breast milk from mothers delivering full term and preterm infants.

Recent advances in the care of low-birth-weight and preterm neonates have stimulated research into the best dietetic program to improve their survival and short/long term outcome. Some components of human milk that cannot be included in artificial formulas may be critical for survival. Of these, immunoglobulins are important, and in particular secretory immunoglobulins A (sIgA). The concentration of secretory IgA was measured by immunoblotting (an immunoelectrophoretic technique having high specificity and reliability) in milk from mothers delivering at term (TM) or prematurely (PM). In both groups, IgA concentrations were high very early on but quickly decreased during the first week of lactation. The early IgA mean concentration was higher in PM than in TM but, because of high variability in PM milk, the difference rarely reached statistical significance. This variability during lactation reflects the important role of human milk in supplying immunological factors to cope with the gastrointestinal absorption of high molecular weight proteins in the first days of life. Immunological protection is particularly critical for a preterm baby, so it is important to promote feeding with its own mothers milk if possible, paying strict attention to the timing of milk collection.

Leukotriene A4, and not leukotriene B4, is the main 5-lipoxygenase metabolite released by bovine leukocytes.

The production of leukotriene A4 (LTA4)-derived metabolites, analysed by RP-HPLC, was studied in purified bovine polymorphonuclear leukocyte (PMNL) preparations and in PMNL-platelet coincubations after challenge with the calcium ionophore A23187. The results obtained show that in bovine PMNL LTB4 represents the main LTA4 metabolite. When washed platelets were added to PMNL, LTC4 was the main enzymatic metabolite observed, indicating a substantial transfer of PMNL-derived LTA4 to platelets. The synthesis of LTC4 was accompanied by a significant decrease in LTB4, suggesting that a quota of the LTB4 synthesized in PMNL preparations is the result of transcellular metabolism of released LTA4 by neighbouring PMNL. Reduction of PMNL-PMNL interactions through dilution of cell incubates allowed us to estimate that most of the leukotriene A4 synthesized by PMNL is indeed released from the cell. LTA4, and not LTB4, represents the main 5-lipoxygenase metabolite released by bovine PMNL.

Cloning and characterization of a rabbit ortholog of human Galpha16 and mouse G(alpha)15.

A cDNA was cloned from a rabbit spleen cDNA library which encoded a G-protein alpha subunit peptide of 374 amino acids, that at the peptide level exhibited 86% and 79% identity with human Galpha16 and mouse G(alpha)15, respectively. The rabbit G(alpha)subunit cDNA was subcloned into a mammalian expression vector and transiently co-transfected into HEK-293 cells along with cDNAs encoding the human C3a, C5a, or nociceptin/orphanin FQ receptors. In all three cases the rabbit G alpha subunit behaved similarly to G(alpha)15 or G(alpha)16 and effectively coupled the transfected receptors to intracellular calcium mobilization pathways. By nucleotide sequence homology and functional activity the rabbit G(alpha) subunit appears to be the ortholog of human G(alpha)16 and mouse G(alpha)15.

Consensus on diuresis renography for investigating the dilated upper urinary tract. Radionuclides in Nephrourology Group. Consensus Committee on Diuresis Renography.

UNLABELLED: There is great variation in technique and interpretation of diuresis renography between different establishments. METHODS: To address this problem, an International Consensus Committee was appointed by the Ninth International Symposium on Radionuclides in Nephrourology in 1994. RESULTS: The final document was produced and addressed: objectives, equipment, data acquisition, choice of radiopharmaceutical, patient preparation, position, dosage of furosemide, timing of furosemide, role of bladder catheter, duration of study, pediatric considerations, evaluation of the furosemide response, interpretation, and conclusion. CONCLUSION: The report presents a standardized approach to diuresis renography that, if adopted, will improve reproducibility between centers, discourage unacceptable practice and stimulate further discussion between nuclear medicine and urology health care professionals who treat patients with dilated and obstructed upper urinary tracts.

The rabbit motilin receptor: molecular characterisation and pharmacology.

Following identification of the human motilin receptor, we isolated the rabbit orthologue by PCR amplification and found this to be 85% identical to the open reading frame of the human receptor. The protein encoded was 84% identical to the human polypeptide. In HEK293T cells transfected with the rabbit receptor, motilin concentration-dependently increased intracellular calcium mobilisation (pEC50=9.25). After transfection with Go1alpha, motilin similarly stimulated [35S]GTPgammaS binding (pEC50=8.87). Using both systems, similar values were obtained with the human receptor, with rank-order potencies of motilin=[Nle13]-motilin>erythromycin; ghrelin was ineffective. In circular muscle preparations of rabbit gastric antrum, [Nle13]-motilin 0.1-30 nM concentration-dependently increased the amplitude of electrically-evoked, neuronally-mediated contractions (pEC50=8.3); higher concentrations increased the muscle tension (30-3000 nM). Both responses to [Nle13]-motilin faded rapidly during its continual presence. Rat or human ghrelin 0.01-10 microM were without activity. Erythromycin 30-3000 nM and 10 microM, respectively, increased neuronal activity and muscle tension in rabbit stomach. Unlike [Nle13]-motilin, the increase in neuronal activity did not fade during continual presence of submaximally-effective concentrations of erythromycin; some fade was observed at higher concentrations. We conclude that the pharmacology of the rabbit motilin receptor is similar to the human orthologue and, when expressed as a recombinant, comparable to the native receptor. However, in terms of their ability to increase neuronal activity in rabbit stomach, [Nle13]-motilin and erythromycin are distinguished by different response kinetics, reflecting different rates of ligand degradation and/or interaction with the receptor.

Molecular and pharmacological characterization of a functional tachykinin NK3 receptor cloned from the rabbit iris sphincter muscle.

1. A functional tachykinin NK3 receptor was cloned from the rabbit iris sphincter muscle and its distribution investigated in ocular tissues. 2. Standard polymerase chain reaction (PCR) techniques were used to clone a full length rabbit NK3 receptor cDNA consisting of 1404 nucleotides. This cDNA encoded a protein of 467 amino acids with 91 and 87% homology to the human and rat NK3 receptors respectively. 3. In CHO-K1 cells transiently expressing the recombinant rabbit NK3 receptor, the relative order of potency of NKB>>NKA>/=SP to displace [125I]-[MePhe7]-NKB binding and to increase intracellular calcium, together with the high affinity of NK3 selective agonists (e.g. senktide, [MePhe7]-NKB) and antagonists (e.g. SR 142801, SB 223412) in both assays was consistent with NK3 receptor pharmacology. In binding and functional experiments, agonist concentration response curves were shallow (0.7 - 0.8), suggesting the possibility of multiple affinity states of the receptor. 4. Quantitative real time PCR analysis revealed highest expression of rabbit NK3 receptor mRNA in iris sphincter muscle, lower expression in retina and iris dilator muscle, and no expression in lens and cornea. In situ hybridization histochemistry revealed discrete specific localization of NK3 receptor mRNA in the iris muscle and associated ciliary processes. Discrete specific labelling of NK3 receptors with the selective NK3 receptor agonist [125I]-[MePhe7]-NKB was also observed in the ciliary processes using autoradiography. 5. Our study reveals a high molecular similarity between rabbit and human NK3 receptor mRNAs, as predicted from previous pharmacological studies, and provide the first evidence that NK3 receptors are precisely located on ciliary processes in the rabbit eye. In addition, there could be two affinity states of the receptor which may correspond to the typical and 'atypical' NK3 receptor subtypes previously reported.

The human GABA(B1b) and GABA(B2) heterodimeric recombinant receptor shows low sensitivity to phaclofen and saclofen.

1. The aim of this study was to characterize the pharmacological profile of the GABA(B1)/GABA(B2) heterodimeric receptor expressed in Chinese hamster ovary (CHO) cells. We have compared receptor binding affinity and functional activity for a series of agonists and antagonists. 2. The chimeric G-protein, G(qi5), was used to couple receptor activation to increases in intracellular calcium for functional studies on the Fluorimetric Imaging Plate Reader (FLIPR), using a stable GABA(B1)/GABA(B2)/G(qi5) CHO cell line. [(3)H]-CGP-54626 was used in radioligand binding studies in membranes prepared from the same cell line. 3. The pharmacological profile of the recombinant GABA(B1/B2) receptor was consistent with that of native GABA(B) receptors in that it was activated by GABA and baclofen and inhibited by CGP-54626A and SCH 50911. 4. Unlike native receptors, the GABA(B1)/GABA(B2)/G(qi5) response was not inhibited by high microMolar concentration of phaclofen, saclofen or CGP 35348. 5. This raises the possibility that the GABA(B1)/GABA(B2)/G(qi5) recombinant receptor may represent the previously described GABA(B) receptor subtype which is relatively resistant to inhibition by phaclofen.

Molecular cloning and characterisation of a novel GABAB-related G-protein coupled receptor.

Using a homology-based bioinformatics approach we have analysed human genomic sequence and identified the human and rodent orthologues of a novel putative seven transmembrane G protein coupled receptor, termed GABA(BL). The amino acid sequence homology of these cDNAs compared to GABA(B1) and GABA(B2) led us to postulate that GABA(BL) was a putative novel GABA(B) receptor subunit. The C-terminal sequence of GABA(BL) contained a putative coiled-coil domain, di-leucine and several RXR(R) ER retention motifs, all of which have been shown to be critical in GABA(B) receptor subunit function. In addition, the distribution of GABA(BL) in the central nervous system was reminiscent of that of the other known GABA(B) subunits. However, we were unable to detect receptor function in response to any GABA(B) ligands when GABA(BL) was expressed in isolation or in the presence of either GABA(B1) or GABA(B2). Therefore, if GABA(BL) is indeed a GABA(B) receptor subunit, its partner is a potentially novel receptor subunit or chaperone protein which has yet to be identified.

Effects of similarity of location and temporal intensity pattern of conditioned and unconditioned stimuli on the acquisition of conditioned suppresion in rats.

This article two studies that examined the effect of similarity of location and temporal intensity patterns of the conditioned stimulus (CS) and the unconditioned stimulus (UCS) on the acquision of conditioned suppression. Experiment 1 showed that greater suppression occurs during a stimulus generalization test when the tested stimulus has a location and temporal intensity pattern that is similar to the previously presented UCS. Experiment 2 showed that conditioned suppression is acquired more rapidly if the CS and UCS have similar locations and temporal intensity patterns. These similarity factors were interpreted as forms of event covariation, which aided in associative formation. Several applications of these data are discussed.

A new remote-controlled endoscope positioning system for endoscopic solo surgery. The FIPS endoarm.

In the field of endoscopic solo surgery, the assistance received by the surgeon from ergonomical positioning devices is extremely important. They aid in both the retracting of instruments and the positioning of the endoscope. However, passive systems derived from open surgery have not proved satisfactory. Therefore, we set out to develop a remote-controlled arm capable of moving a rigid endoscope with about four degrees of freedom, while maintaining an invariant point of constraint motion coincident with the trocar puncture site through the abdominal wall. The system is driven by means of speaker-independent voice control or a finger-ring joystick clipped onto the instrument shaft close to the handle. When the joystick is used, the motion of the endoscope is controlled by the fingertip of the operating surgeon, which is inserted into the small ring of the controller in such a way as to make the motion of the fingertip correspond directly to the motion of the tip of the endoscope. A study was performed to compare the two different interfaces available for the system. With both interfaces, the guiding system allows for transparent and intuitive operation. Its set-up is easy; it is safe and reliable to use during the intervention; and it is faster than human assistance. With its improved ergonomy, this new generation of remote-controlled endoscope positioning system represents a further step toward the diffusion of solo surgery techniques in minimally invasive therapy. In our opinion, this prototype creates a valid compromise between human and robotic control of rigid endoscopes.

Carcinosarcoma of the lung: a case report.

The clinical and pathological data of a single case of carcinosarcoma of the lung observed in the period from 1978 to 1998 were reviewed. The diagnosis was based on immunohistochemical examination of the surgical specimen after a lower left lobectomy. The patient was given adjuvant chemotherapy. The local recurrence showed only sarcomatous features. The characteristics of this rare tumor are discussed in this case report.

Extension of lymph node dissection and survival in primary gastric cancer.

The histories of 429 patients who underwent surgery for primary gastric cancer at our ward from January 1970 to December 1985, were reviewed. All patients underwent surgery: potentially curative surgery, 54.8%, non-curative resection, 18.2%; palliative surgery, 27%. Nodal status was as follows: N0, 28%; N1, 17.7%; N2, 44.5%; N3, 9.8%. The incidence of N0 cases was significantly increased in Stage T1 and T2 disease compared to Stage T3 and T4 lesions (p < 0.001). In Stage T3 and T4 patients the incidence of distant metastases increased if lymph node involvement was also present (p < 0.005). In patients without nodal metastases 5-year survival was 70% (median survival: 60+ months) whereas, in patients with lymph node involvement survival was 32% (median survival: 24 months) (p < 0.001). Our data suggest that elective extensive lymph node dissection (R2) is indicated in all patients because survival is improved by this procedure. We recommend R3 lymph node dissection only in macroscopic N3 node involvement patients.

[Diagnostic peritoneal lavage in abdominal traumatology. Review of the literature]. / Il lavaggio peritoneale diagnostico nella traumatologia addominale. Revisione della letteratura.

This paper presents an exhaustive review of the literature on diagnostic peritoneal lavage (indications, contraindications, technique and results), since nowadays this constitutes the simplest and most reliable diagnostic procedure for internal and open abdominal trauma.

[Nuclear DNA content in tumors of the colon and rectum. Relationship between ploidy and site of the neoplasm]. / Contenuto nucleare di DNA nei tumori del colon-retto. Rapporto tra ploidia e sede della neoplasia.

The DNA distribution pattern was determined 30 colo-rectal carcinomas. The DNA content in individual cells was measured according to a cytophotometric method based on light transmission measurement of Feulgen-stained nuclei. The DNA non diploid carcinomas were significantly more common from the left (descending and sigmoid) colon and rectum (P = 0.0002). No relationship existed between DNA pattern and age, sex, Dukes' stage, tumor grade.

[Positioning systems for endoscopic solo surgery]. / Sistemi di posizionamento per solo chirurgia endoscopica.

BACKGROUND: Endoscopic surgery has acquired undisputed importance in the field of both general and specialised surgery. The introduction of robotic technology in surgery has recently led to the development of new positioning systems for endoscopic surgery. These allow direct control of the endoscopic procedures by the surgeon, whose vision currently depends on the assistant in charge of positioning the optic camera in compliance with his wishes. METHODS: We experimented different positioning systems for optics and rigid endoscopic instruments for laparoscopy, some of which were our own design. Over 400 cholecystectomies were carried out by six different surgeons on phantoms containing animal organs. The experimental systems were AESOP (Computer Motion, USA), with both foot-pedal and voice control, ENDOASSIST (Armstrong Healthcare Co. UK), controlled by a device worn by the surgeon, FIPS Endoarm (Karlsruhe Research Centre, Germany), controlled by a joystick and voice, and the passive TISKA Endoarm system (Karlsruhe Research Centre, Germany). Combinations of two systems were compared, using one to position the optic and one to position the retractor instrument. RESULTS: Phantom tests, which are preferable owing to constant conditions, showed the feasibility of experiments in Solo Surgery conditions and highlighted the advantages and drawbacks of the various systems. In particular, the surgeons appreciated the intuitive use of the TISKA Endoarm system as a positioner for the retractor instrument and the optics, in spite of the fact that it was only a passive movement apparatus. Among the remote-control systems tested as an optics positioner, FIPS Endoarm controlled by a joystick was particularly intuitive and produced the best results in terms of time taken to complete the procedure. The time taken was even shorter than that in a large control group with human assistance. CONCLUSIONS: In our experience endoscopic Solo Surgery was found to be applicable to clinical practice. This will bring numerous advantages in terms of the precision of surgical procedures and savings in terms of time and human resources, with a consequent reduction of management costs. There is no doubt that this method represents a step forward in the application of technology to surgery.