Inhibiting transthyretin conformational changes that lead to amyloid fibril formation

Insoluble protein fibrils resulting from the self-assembly of a conformational intermediate are implicated as the causative agent in several severe human amyloid diseases, including Alzheimer's disease, familial amyloid polyneuropathy, and senile systemic amyloidosis. The latter two diseases are associated with transthyretin (TTR) amyloid fibrils, which appear to form in the acidic partial denaturing environment of the lysosome. Here we demonstrate that flufenamic acid (Flu) inhibits the conformational changes of TTR associated with amyloid fibril formation. The crystal structure of TTR complexed with Flu demonstrates that Flu mediates intersubunit hydrophobic interactions and intersubunit hydrogen bonds that stabilize the normal tetrameric fold of TTR. A small-molecule inhibitor that stabilizes the normal conformation of a protein is desirable as a possible approach to treat amyloid diseases. Molecules such as Flu also provide the means to rigorously test the amyloid hypothesis, i.e., the apparent causative role of amyloid fibrils in amyloid disease.     

Pathogenesis of amniotic fluid embolism

The reason for existant of amniotic fluid embolism is the disposition, which will be favoured by the overaction of the sympathic system, by age and the number of pregnancy. It happens especially by premature rupture of membrane, pathologic contents in the amniotic fluid, high pressure in the uterus, bad uterine muscel with laceration and with opened endymyometric veins. Pathophysiologic connection between the stages of amniotic fluid embolism are discussed in detail.     

Intraperitoneal pressure: ascitic fluid and splanchnic vascular pressures, and their role in prevention and formation of ascites

Seventeen patients with ascites due to cirrhosis underwent hepatic venous catheterization and pressure measurement in the ascitic fluid. Intraperitoneal fluid hydrostatic pressure (IFP) ranged 3.5-22, mean 11.2 mm Hg, and correlated closely to the pressure in the inferior vena cava (r = 0.97, P < 0.001), which was on average 1.8 mmHg above that of ascitic fluid (P < 0.005). Wedged hepatic venous pressure (WHVP) (range 19-43, mean 32 mmHg) correlated directly to IFP (0.89, P < 0.001) and was significantly higher than that of ten cirrhotic patients without ascites (range 12-27, mean 20 mmHg, P < 0.005). After diuretic therapy WHVP decreased to an average of 20 mmHg. Mean plasma colloid osmotic pressures were 20 mmHg (range 18-24 mmHg)( and 23 mmHg (range 19-29 mmHg) in patients with and without ascites, the values being significantly different (P < 0.05). Colloid osmotic pressure of ascitic fluid ranged 1-14, mean 4.9 mmHg. Mean ratio between albumin concentration in ascitic fluid and plasma was 0.31 (range 0.12-0.77). In five pigs portal venous pressure (PVP) increased during infusion of fluid into the peritoneal cavity. The increase in PVP was smaller than that of IFP (P < 0.02), indicating that ascitic fluid stems the pressures in the splanchnic venous vascular bed up to a higher level, but that the transmural hydrostatic pressure difference decreases simultaneously. The results are discussed in relation to the local 'oedema-preventing' mechanisms: (a) increased interstitial hydrostatic fluid pressure, (b) decreased interstitial fluid colloid osmotic pressure, (c) increased lymph flow, and it is concluded that the peritoneal space can be considered as a special part of the interstitium in which IFP is considered to play an important role in regulation of ascitic fluid.     

Acquired conditions that lead to osteoarthritis in the dog

There are many acquired arthopathies that will result in some degree of osteoarthritis, even after proper management. Once the articular cartilage is damaged, it is unlikely that the architecture of the original cartilage surface will return to the normal conditions that existed prior to injury. The purpose of timely and meticulous management of traumatic joint events is to stop the progression of osteoarthritic development. When dealing with articular fractures or other forms of trauma to articular cartilage, three important principles to remember are anatomic reduction of the articular surfaces, stable fixation, and limited weight bearing on the affected limb as soon as possible after surgery. Even after strict adherence to these principles, the pet owner should always be warned that the animal will develop some degree of osteoarthritis in the affected joint at some future time; at that time, chronic medical management may be indicated.     

Congenital conditions that lead to osteoarthritis in the dog

Several congenital arthropathies exist in the canine that can lead to the development of degenerative joint disease. Nevertheless, early diagnosis and treatment generally will afford the patient a favorable prognosis for limb function. There is still a great need to develop controlled studies to evaluate the long-term efficacy of many of the surgical and medical treatment modalities that are currently available and will be made available in the future to treat the patient with a congenital arthropathy.     

Potential mechanisms of metabolic imprinting that lead to chronic disease

This review synthesizes a subset of human epidemiologic and experimental animal studies that suggest that early nutrition affects susceptibility to chronic diseases in adulthood. These studies provide evidence that biological mechanisms may exist to "memorize" the metabolic effects of early nutritional environments. However, hypothesis-driven investigations of potential mechanisms have been scant. Thus, our understanding of the biology underlying metabolic imprinting is incomplete. A working definition of metabolic imprinting is proposed, emphasizing the adaptive nature and limited ontogenic window of the mechanisms putatively responsible for these relations. Five specific candidate mechanisms of metabolic imprinting are elaborated: 1) induced variations in organ structure, 2) alterations in cell number, 3) clonal selection, 4) metabolic differentiation, and 5) hepatocyte polyploidization. Last, experimental approaches for probing potential mechanisms with animal models are discussed.     

A quantitative measurement of peritoneal drainage in malignant ascites

With a two-compartment model, a method is described for quantitative determination of peritoneal drainage rates in malignant ascites. Data on twenty-four patients are presented and comparison is made with the qualitative assessment of the integrity of diaphragmatic and mediastinal lymphatics on the basis of lymphoscintigrams. It is concluded that flow rates less than 50 ml/hour are usually associated with abnormalities of the diaphragmatic and mediastinal lymphatics, indicating that tumor permeation of these structures is a significant contributing factor to persistent, intractable ascites in patients with malignant diseases.     

Clinical evaluation of polyamine assay in diagnosis of malignant ascites

Polyamine levels were determined by precolumn derivation RH-HPLC in 68 patients with ascites including 21 malignant ascites, 28 liver cirrhosis and 19 tuberculosis. The total polyamine, especially the contents of cadaverine (CA) and spermidine (SPD) in malignant ascites were significantly higher than that in cirrhosis and tuberculosis. Among them CA levels in malignant ascites and cirrhotic and tubercular ascitic fluid were 2.26 +/- 1.83 nmol.ml-1, 0.75 +/- 0.52 nmol.ml-1 and 0.76 +/- 0.47 nmol.ml-1 respectively, demonstrated the most obvious difference. Using 1.0 nmol.ml-1 as a threshold cut off point to differentiate the malignant ascites from non-malignant ascites, the from non-malignant ascites, the sensitivity, specificity and accuracy reached 76. 2%, 83.0% and 80.9% respectively. The results suggest that polymine level in ascites might be regard as one of the cancer-associated markers.     

Tryptophan synthase mutations that alter cofactor chemistry lead to mechanism-based inactivation

Mutations in the pyridoxal phosphate binding site of the tryptophan synthase beta subunit (S377D and S377E) alter cofactor chemistry [Jhee, K.-H., et al. (1998) J. Biol. Chem. 273, 11417-11422]. We now report that the S377D, S377E, and S377A beta2 subunits form alpha2 beta2 complexes with the alpha subunit and activate the alpha subunit-catalyzed cleavage of indole 3-glycerol phosphate. The apparent Kd for dissociation of the alpha and beta subunits is unaffected by the S377A mutation but is increased up to 500-fold by the S377D and S377E mutations. Although the three mutant alpha2 beta2 complexes exhibit very low activities in beta elimination and beta replacement reactions catalyzed at the beta site in the presence of Na+, the activities and spectroscopic properties of the S377A alpha2 beta2 complex are partially repaired by addition of Cs+. The S377D and S377E alpha2 beta2 complexes, unlike the wild-type and S377A alpha2 beta2 complexes and the mutant beta2 subunits, undergo irreversible substrate-induced inactivation by L-serine or by beta-chloro-L-alanine. The rates of inactivation (kinact) are similar to the rates of catalysis (kcat). The partition ratios are very low (kcat/kinact = 0.25-3) and are affected by alpha subunit ligands and monovalent cations. The inactivation product released by alkali was shown by HPLC and by fluorescence, absorption, and mass spectroscopy to be identical to a compound previously synthesized from pyridoxal phosphate and pyruvate. We suggest that alterations in the cofactor chemistry that result from the engineered Asp377 in the active site of the beta subunit may promote the mechanism-based inactivation.     

Pleural fluid accumulation due to intra-abdominal endometriosis: a case report and review of the literature

A case is presented of massive ascites and right sided pleural effusion caused by endometriosis. The final diagnosis was not made for a considerable time. Massive ascites and a right sided pleural effusion caused by endometriosis is rare, with fewer than 10 reports in the literature worldwide. Physicians should be aware of this potentially tentially treatable cause, having excluded other possibilities such as malignancy and tuberculosis.     

Microsaccadic response to visual events that are invisible to the superior colliculus.

Even when people think their eyes are still, tiny fixational eye movements, called microsaccades, occur at a rate of –1 Hz. Whenever a new (and potentially dangerous) event takes place in the visual field, the microsaccadic frequency is at first inhibited and then is followed by a rebound before the frequency returns to baseline. It has been suggested that this inhibition-rebound response is a type of oculomotor reflex mediated by the superior colliculus (SC), a midbrain structure involved in saccade programming. The present study investigated microsaccadic responses to visual events that were invisible to the SC; the authors recorded microsaccadic responses to visual oddballs when the latter were equiluminant with respect to the standard stimuli and when both oddballs and standards were equiluminant with respect to the background. Results showed that microsaccadic responses to oddballs and to standards were virtually identical both when the stimuli were visible to the SC and when they were invisible to it. Although the SC may be the generator of microsaccades, this research suggests that the specific fixational oculomotor activity in response to visual events can be controlled by other brain centers. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Translation of Sindbis virus mRNA: analysis of sequences downstream of the initiating AUG codon that enhance translation

Alphaviruses, particularly Sinbis virus and Semliki Forest virus, are proving to be useful vectors for the expression of heterologous genes. In infected cells, these self-replicating vectors (replicons) transcribe a subgenomic mRNA that codes for a heterologous protein instead of the structural proteins. We reported recently that translation of the reporter gene lacZ is enhanced 10-fold when the coding sequences of this gene are fused downstream of and in frame with the 5' half of the capsid gene (I. Frolov and S. Schlesinger, J. Virol. 68:8111-8117, 1994). The enhancing sequences, located downstream of the AUG codon that initiates translation of the capsid protein, have a predicted hairpin structure. We have mutated this region by making changes in the codons which do not affect the protein sequence but should destabilize the putative hairpin structure. These changes caused a decrease in the accumulation of the capsid-beta-galactosidase fusion protein. When these alterations were inserted into the capsid gene in the context of the intact Sindbis virus genome, they led to a decrease in the rate of virus formation but did not affect the final yield. We also altered the original sequence to one that has 12 contiguous G.C base pairs and should form a stable hairpin. The new sequence was essentially as effective as the original had been in enhancement of translation and in the rate of virus formation. The position of the predicted hairpin structure is important for its function; an insertion of 9 nucleotides or a deletion of 9 nucleotides decreased the level of translation. The insertion of a hairpin structure at a particular location downstream of the initiating AUG appears to be a way that alphaviruses have evolved to enhance translation of their mRNA, and, as a consequence, they produce high levels of the structural proteins which are needed for virus assembly. This high level of translation requires an intracellular environment in which host cell protein synthesis is inhibited.     

Differential inhibition of fluid accumulation and tumor growth in two mouse ascites tumors by an antivascular endothelial growth factor/permeability factor neutralizing antibody

In the accompanying paper (Luo et al., Cancer Res., 58: 2652-2660, 1998), we demonstrated that vascular endothelial growth factor (VEGF), also designated vascular permeability factor (VPF), significantly accumulated in all mouse malignant ascites tested, suggesting its fundamental role in ascites tumors. Removal of VEGF may inhibit the development of ascites tumors. In this study, using a goat antimouse VEGF-neutralizing antibody, we tested this hypothesis with two well-defined syngeneic mouse ascites tumors: MM2 breast adenocarcinoma and OG/Gardner lymphoma 6C3HED (expressing moderate and low levels of VEGF, respectively). This antibody significantly inhibited MM2 and OG cell-free ascites fluid-induced hyperpermeability of mouse peritoneal microvessels and in vitro endothelial cell growth. Mice bearing tumors were administered i.p. daily with the antibody or normal goat IgG as controls for 8 days, at doses of 20-fold (for MM2-bearing mice) or 40-fold (for OG-bearing mice) the estimated amounts of VEGF that kinetically accumulated in the ascites fluid after the tumor inoculation. The average volume of ascites fluid, number of tumor cells and leaked RBCs, and the peritoneal microvessel permeability in MM2-bearing mice that received the antibody treatment were significantly lower than those in the matched controls (P < 0.01). Unexpectedly, OG-bearing mice did not show satisfactory response to the anti-VEGF treatment. This discrepancy was not likely due to inadequate doses or different host immune responses, but it was quite possibly to the different characteristics of MM2 carcinoma and OG lymphoma tumors, the latter being strongly invasive, and/or the existence of an inflammatory mediator(s), such as bradykinin or cytokine(s) other than VEGF. In summary, our results directly demonstrated, for the first time, differential roles for VEGF in ascites tumors in vivo and suggest the potential of VEGF inhibition as a specific therapy for ascites tumors of carcinoma origin, which are the major cause of the malignant ascites in adult humans.     

Denervation of pigment cells lead to a receptor that is ultrasensitive to melatonin and noradrenaline

Pigment granule aggregation and dispersal can be studied in the melanophores of isolated scales from the cuckoo wrasse (Labrus ossifagus L.). Stimulation of a melanophore alpha2-adrenoceptor or the sympathetic nerve innervating the cell causes pigment aggregation. When the stimulation ceases, the pigment granules disperse throughout the cell. Studying this migration has been a useful tool in pharmacological research, particularly in investigations of the alpha2-adrenoceptor. Denervation of melanophores creates a receptor that is ultrasensitive to noradrenaline and melatonin. After three to four weeks of isolation, the denervated melanophores exhibit a 10(9)-fold increase in sensitivity. The efficacy of melatonin is increased from a negligible pigment-aggregation ability to the level of a full agonist. The melatonin-induced aggregation can, however, be counteracted by the alpha2-adrenoceptor antagonist yohimbine, but not by alpha1-adrenoceptor antagonist prazosin, indicating that the ultrasensitive receptor possesses alpha2-adrenoceptor features. Consequently, we conclude that the ultrasensitive receptor may represent an alpha2-adrenoceptor that has, due to denervation of the melanophore, become sensitive to melatonin.     

Mutations in nucleolar proteins lead to nucleolar accumulation of polyA+ RNA in Saccharomyces cerevisiae

Synthesis of mRNA and rRNA occur in the chromatin-rich nucleoplasm and the nucleolus, respectively. Nevertheless, we here report that a Saccharomyces cerevisiae gene, MTR3, previously implicated in mRNA transport, codes for a novel essential 28-kDa nucleolar protein. Moreover, in mtr3-1 the accumulated polyA+ RNA actually colocalizes with nucleolar antigens, the nucleolus becomes somewhat disorganized, and rRNA synthesis and processing are inhibited. A strain with a ts conditional mutation in RNA polymerase I also shows nucleolar accumulation of polyA+ RNA, whereas strains with mutations in the nucleolar protein Nop1p do not. Thus, in several mutant backgrounds, when mRNA cannot be exported i concentrates in the nucleolus. mRNA may normally encounter nucleolar components before export and proteins such as Mtr3p may be critical for export of both mRNA and ribosomal subunits.     

Antibiotic concentrations in ascitic fluid of patients with ascites and bacterial peritonitis

Thirty-six paired specimens of serum and ascitic fluid from 21 patients with peritonitis and ascites, most with sponetaneous bacterial peritonitis and alcoholic cirrhosis, were assayed for antibiotic content. Antibiotics assayed and number of determinations were gentamicin, 14; tobramycin, 7; ampicillin, 5; clindamycin, 3; penicillin G, 2; cephalothin, 2; chloramphenico, 2; and cefazolin, 1. In 31 pared specimens the ascitic fluid antibiotic concentration was about one half or more of the simultaneous serum level and in 17 assays exceeded 90% of the serum level. All antibiotics studied penetrated ascitic fluid equally well. Clinical response to antibiotic therapy was good in 12 of 16 patients with culture-proven bacterial peritonitis. Antibiotic levels in ascitic fluid exceeded the minimal inhibitory concentration of the infecting organisms in all but one patient who responded. Direct intraperitoneal instillation of antibiotics does not appear to be necessary routinely; however, there may be an initial lag of several hours before antibiotic concentrations is ascites achieve therapeutic levels.     

Initial stages of the formation of lead oxide phases

Metal lead films are studied by electron diffraction during heating depending on temperature T and oxygen partial pressure p _O2 in the gas phase. It is found that the formation of oxide phases begins from the α-PbO₂ modification. As p _O2 increases or T increases, intermediate oxides nPbO₂ · mPbO and Pb₃O₄ form and then transform into the β-PbO modification.     

Role of vascular endothelial growth factor in ovarian cancer: inhibition of ascites formation by immunoneutralization

Ovarian cancer is characterized by the rapid growth of solid intraperitoneal tumors and large volumes of ascitic fluid. Vascular endothelial growth factor (VEGF) augments tumor growth by inducing neovascularization and may stimulate ascites formation by increasing vascular permeability. We examined the role of VEGF in ovarian carcinoma using in vivo models in which intraperitoneal or subcutaneous tumors were induced in immunodeficient mice using the human ovarian carcinoma cell line SKOV-3. After tumor engraftment (7 to 10 days), some mice were treated with a function-blocking VEGF antibody (A4.6.1) specific for human VEGF. A4.6.1 significantly (P < 0.05) inhibited subcutaneous SKOV-3 tumor growth compared with controls. However, tumor growth resumed when A4.6.1 treatment was discontinued. In mice bearing intraperitoneal tumors (IP mice), ascites production and intraperitoneal carcinomatosis were detected 3 to 7 weeks after SKOV-3 inoculation. Importantly, A4.6.1 completely inhibited ascites production in IP mice, although it only partially inhibited intraperitoneal tumor growth. Tumor burden was variable in A4.6.1-treated IP mice; some had minimal tumor, whereas in others tumor burden was similar to that of controls. When A4.6.1 treatment was stopped, IP mice rapidly (within 2 weeks) developed ascites and became cachectic. These data suggest that in ovarian cancer, tumor-derived VEGF is obligatory for ascites formation but not for intraperitoneal tumor growth. Neutralization of VEGF activity may have clinical application in inhibiting malignant ascites formation in ovarian cancer.