Usefulness of continuous air tonometry for evaluation of splanchnic perfusion during cardiopulmonary bypass

Although gastric mucosal tonometry has been reported as a useful method to assess splanchnic perfusion during cardiovascular surgery, the conventional discontinuous method of tonometry (saline tonometry) was cumbersome and prone to systematic errors. A new automated system of air tonometry (Tonocap; Datex Ohmeda, Helsinki, Finland) allows for frequent (every 10 minutes) measurement of gastric regional CO2 (PrCO2) and may be more suitable as a monitoring system in cardiac patients. We evaluated the usefulness of continuous air tonometry as a marker of splanchnic perfusion during cardiopulmonary bypass (CPB). In 19 patients (53-79 years, mean 63 years) who underwent cardiovascular surgery under standard CPB with mild hypothermia (32 degrees C) from January 2001 to May 2002, the PrCO2 and calculated intramucosal pH (pHi) of gastric tonometry was monitored using Tonocap, and their relation to postoperative visceral organ function was evaluated. The pHi significantly increased after initiation of CPB from 7.32 +/- 0.07 to 7.43 +/- 0.10 (p < 0.05) and then consistently decreased in all patients to 7.39 +/- 0.09 at the end of CPB. The value of PrCO2 significantly (p < 0.01) correlated with the value of pHi. The lowest value of pHi during CPB was significantly related to blood urea nitrogen (r = -0.75, p < 0.05), serum creatinine (r = -0.78, p < 0.05), creatinine clearance (r = 0.68, p < 0.05) on postoperative day 1, and blood urea nitrogen (r = -0.84, p < 0.01) on day 3. In contrast, arterial blood lactate level, venous oxygen saturation, and routinely measured hemodynamics (e.g., pump flow, arterial pressure) during CPB were unrelated to the postoperative visceral organ function. These results suggest that continuous monitoring of gastric regional CO2 and pHi by air tonometry system is useful for the evaluation of splanchnic perfusion during CPB and may contribute to improve CPB technique by allowing the early detection of visceral malperfusion.     

Lung Infection Caused by Gordona aurantiaca (Rhodococcus aurantiacus)

We report that Gordona aurantiaca (Rhodococcus aurantiacus) caused a lung infection in humans. The case of a 50-year-old male farmer is shown. This is the first report of infection by this organism.     

Behaviour of permeation and separation for aqueous organic acid solutions through poly(vinyl chloride) and poly[(vinyl chloride)-co-(vinyl acetate)] membranes

The permeation and separation characteristics of poly(vinyl chloride) (PVC) and poly[(vinyl chloride)-co-(vinyl acetate)] (poly(VC-co-VAc)) membranes were investigated for aqueous organic acid solutions by pervaporation and evapomeation. The PVC membrane preferentially incorporates organic acids and predominantly permeates water from aqueous organic acid solutions. Water permselectivities of these aqueous solutions through the PVC membrane are significantly dependent on high diffusivity of water across the membrane. It was found that the permeation rate increases and the separation factor for the water permselectivity decreases with increasing vinyl acetate (VAc) content in the poly(VC-co-VAc) membrane. Preferential solubility of acetic acid into the poly(VC-co-VAc) membrane increases with the VAc content. This result was explained by a strong affinity between acetic acid and the VAc unit in the poly(VC-co-VAc) membrane.     

Two closely related ubiquitin C-terminal hydrolase isozymes function as reciprocal modulators of germ cell apoptosis in cryptorchid testis

The experimentally induced cryptorchid mouse model is useful for elucidating the in vivo molecular mechanism of germ cell apoptosis. Apoptosis, in general, is thought to be partly regulated by the ubiquitin-proteasome system. Here, we analyzed the function of two closely related members of the ubiquitin C-terminal hydrolase (UCH) family in testicular germ cell apoptosis experimentally induced by cryptorchidism. The two enzymes, UCH-L1 and UCH-L3, deubiquitinate ubiquitin-protein conjugates and control the cellular balance of ubiquitin. The testes of gracile axonal dystrophy (gad) mice, which lack UCH-L1, were resistant to cryptorchid stress-related injury and had reduced ubiquitin levels. The level of both anti-apoptotic (Bcl-2 family and XIAP) and prosurvival (pCREB and BDNF) proteins was significantly higher in gad mice after cryptorchid stress. In contrast, Uchl3 knockout mice showed profound testicular atrophy and apoptotic germ cell loss after cryptorchid injury. Ubiquitin level was not significantly different between wild-type and Uchl3 knockout mice, whereas the levels of Nedd8 and the apoptotic proteins p53, Bax, and caspase3 were elevated in Uchl3 knockout mice. These results demonstrate that UCH-L1 and UCH-L3 function differentially to regulate the cellular levels of anti-apoptotic, prosurvival, and apoptotic proteins during testicular germ cell apoptosis.     

En(a-) phenotype in a Japanese blood donor

The first Japanese En(a-) individual (T.N.) was found by screening red cells from 250,000 Japanese blood donors with monoclonal anti-Ena. His serum contained no atypical antibodies and his partial red cell phenotype was M-N-S+s-, although a trypsin- resistant N antigen was detected. His red cells were En(a-) and Wr(b-), as determined by various human and mouse monoclonal antibodies. The absence of glycophorin A (GPA) and the presence of apparently normal glycophorin B (GPB) were demonstrated by immunoblotting with antibodies to the extracellular and cytoplasmic domain of GPA and to epitopes common to GPA and GPB. Sialic acid levels of T.N.'s intact red cells were substantially lower than those of control MN cells. Serologic tests suggested that both of T.N.'s parents were heterozygous for a recessive GPA deficiency gene.     

Dynamic alteration of human β-defensin 2 localization from cytoplasm to intercellular space in psoriatic skin

Defensins are cationic antimicrobial peptides with a broad spectrum. Recently human beta-defensin 2 (hBD-2) has been isolated from psoriatic skin; however, its exact localization and fate have not been fully understood. We studied the distribution pattern of hBD-2 in skin tissues of psoriasis and other inflammatory skin diseases. In the upper spinous and granular layer of psoriasis vulgaris hBD-2 was present in the cytoplasm. In the horny layer the positive signals were in a basket-weave pattern, indicating possible accumulation of hBD-2 in the intercellular space. The similar pattern of hBD-2 distribution was observed in the lesions of nummular eczema and atopic dermatitis. hBD-2 was not detected in the section of normal elbow and knee skin. When isolated psoriatic scales were stained, hBD-2 was detected in a wrapping paper-like distribution pattern surrounding the corneocytes. In horny layer of psoriatic skin hBD-2 was closely associated or colocalized with elafin, which is known to be in extracellular space, as demonstrated by double staining. Western blot analysis using cultured human keratinocytes detected hBD-2 with an expected size in the conditioned medium and in the cell lysates when stimulated with 5% FCS or IL-alpha. These results indicate that hBD-2 was synthesized and remained in cytoplasm in the upper spinous and granular layer, and then secreted into intercellular space in the horny layer. This dynamic change in hBD-2 distribution in epidermis is certainly relevant to function as an innate host defense mechanism against invading micro-organisms.     

Purification of fully activated Clostridium botulinum serotype B toxin for treatment of patients with dystonia

Clostridium botulinum serotype B toxins 12S and 16S were separated by using a beta-lactose gel column at pH 6.0; toxin 12S passed through the column, whereas toxin 16S bound to the column and eluted with lactose. The fully activated neurotoxin was obtained by applying the trypsin-treated 16S toxin on the same column at pH 8.0; the neurotoxin passed through the column, whereas remaining nontoxic components bound to the column. The toxicity of this purified fully activated neurotoxin was retained for a long period by addition of albumin in the preparation.     

Ex vivo localization of the mouse saposin C activation region for acid beta-glucosidase

Saposin C is a biological activator of acid beta-glucosidase (GCase), the lysosomal hydrolase with activity towards glucosylceramide (GC). In addition, saposin C possesses a functional domain that determines the in vitro and ex vivo neuritogenic effects of prosaposin, the precursor of saposins A, B, C, and D. The domains for enzymatic activation and neuritogenic function segregate in vitro, respectively, to the carboxyl- and amino-terminal halves of human and mouse saposin C. A chimeric mouse saposin C(1-8)B(8-28)C(30-80) was created to obliterate the neuritogenic region by substituting amino acids 9-29 of saposin C with amino acids 8-28 of saposin B. This saposin showed normal in vitro enzymatic activation effects toward GCase, but no neuritogenic activity. An altered prosaposin was made to contain the chimeric saposin C region. Expression of this altered or wild-type prosaposin was driven by the PGK-1 promoter as a transgene in prosaposin knock-out mice. In cultured fibroblasts from such mice, expressed saposins localized to the lysosomal compartments. Metabolic lipid labeling using L-[3-(14)C]serine showed retention or clearance of GC in prosaposin deficient or transgene reconstituted cells, respectively. In addition, sulfatide catabolism, that requires saposin B and arylsulfatase, was also normalized in prosaposin KO cells reconstituted with the transgenes. These data show that the transgenic prosaposins were expressed and processed to functional saposins in fibroblasts. These results also show that the enzymatic activation domain is located at carboxyl-terminal half of saposin C and functions only in the context of the general saposin structure.     

Prosaposin: threshold rescue and analysis of the "neuritogenic" region in transgenic mice

Prosaposin is the precursor of four glycoprotein activators (saposins) for lysosomal hydrolases. Intact prosaposin also has lipid transfer properties in vitro as well as neuritogenic effects ex vivo and in vivo. Such "neuritogenic" effects of saposin C were evaluated in vivo using transgenic mice with prosaposin cDNAs having normal (PS-N) or mutated neuritogenic region. The mutant prosaposin cDNA (PS-CBC) encoded a chimeric saposin C that contained the non-neuritogenic sequence of saposin B, but retained acid beta-glucosidase (GCase) activation effects. When driven by the PGK (3-phosphoglycerate kinase) promoter, transgene expression was highest in the cerebrum for any of the transgenes (range from 15% to 42% of wild-type). Low levels were in visceral tissues. Prosaposin knock-out (PS-/-) mice expressing N or CBC transgenes, even at low levels, had delayed onset of neurologic signs and neuropathology, and significant lengthening of life span (from 1.7- to 7-fold) with age dependent partial correction of GlcCer and LacCer accumulation in the brain. Neuropathologic progression and neuronal glycosphingolipid storage were related directly to the transgene expression levels in the brain. Purkinje cell loss was age dependent. Gross brain and neuronal organizations were indistinguishable in PS-/- mice with or without the various transgenes, albeit the phenotype appeared later in the mice with transgenes. These studies show the degree of neuropathologic manifestations in each transgenic line depended on expression level rather than on the nature of the transgene. These studies also show in vivo localization of the GCase activation region to the carboxy terminal half of saposin C and the lack of a significant gross trophic effect of saposin C on CNS organization in vivo.