Targeting Semaphorin 3C in Prostate Cancer With Small Molecules.

Despite the amenability of early-stage prostate cancer to surgery and radiation therapy, locally advanced and metastatic prostate cancer is clinically problematic. Chemical castration is often used as a first-line therapy for advanced disease, but progression to the castration-resistant prostate cancer phase occurs with dependable frequency, largely through mutations to the androgen receptor (AR), aberrant AR signaling, and AR-independent mechanisms, among other causes. Semaphorin 3C (SEMA3C) is a secreted signaling protein that is essential for cardiac and neuronal development and has been shown to be regulated by the AR, to drive epithelial-to-mesenchymal transition and stem features in prostate cells, to activate receptor tyrosine kinases, and to promote cancer progression. Given that SEMA3C is linked to several key aspects of prostate cancer progression, we set out to explore SEMA3C inhibition by small molecules as a prospective cancer therapy. A homology-based SEMA3C protein structure was created, and its interaction with the neuropilin (NRP)-1 receptor was modeled to guide the development of the corresponding disrupting compounds. Experimental screening of 146 in silico‒identified molecules from the National Cancer Institute library led to the discovery of four promising candidates that effectively bind to SEMA3C, inhibit its association with NRP1, and attenuate prostate cancer growth. These findings provide proof of concept for the feasibility of inhibiting SEMA3C with small molecules as a therapeutic approach for prostate cancer.

Feasibility of second hematopoietic stem cell transplantation using reduced-intensity conditioning with fludarabine and melphalan after a failed autologous hematopoietic stem cell transplantation.

This study was performed to determine the feasibility of second hematopoietic stem cell transplantation (HSCT) using reduced-intensity conditioning (RIC) with fludarabine and melphalan in patients with relapsed hematologic malignancies after a prior autologous HSCT. Twelve patients (multiple myeloma [n = 7], non-Hodgkin lymphoma [n = 3], and acute myeloid leukemia [n = 2] received allogeneic HSCT using RIC with fludarabine (25 mg/m(2) for 5 days) and melphalan (140 mg/m(2) for 1 day) after a failed autologous HSCT. The graft-versus-host disease (GVHD) prophylaxis consisted of cyclosporine plus a minidose of methotrexate. All patients achieved a neutrophil and platelet engraftment in a median 13.5 days and 17.5 days, respectively. The transplant-related mortality was 2 patients (16.7%). Grade II-IV acute GVHD and chronic extensive GVHD were noted in 4 (33.3%) and 1 patient (11.1%), respectively. Over a median follow-up duration of 376 days, 5 patients were alive without evidence of disease. The estimated nonrelapse mortality at 1 year was 28.4%. The estimated overall survival rate at 1 year was 58.3%, and the estimated event-free survival rate at 1 year was 41.7%. Allogeneic HSCT using RIC with fludarabine and melphalan appears to be feasible for a second HSCT in patients with relapsed hematologic malignancies after a failed autologous HSCT.

Effect of saliva contamination on bond strength of resin luting cements to dentin.

OBJECTIVES: This study examined the effect of saliva contamination on the microtensile bond strength (microTBS) of resin luting cements to dentin. METHODS: For RelyX ARC (ARC, 3M ESPE), dentin surfaces were etched with 32% phosphoric acid. The subgroups were: ARC-control (uncontaminated), ARC-I (saliva contamination, blot-dried), ARC-II (saliva contamination, rinse, blot-dried) and ARC-III (saliva contamination, rinse, re-etch, rinse, blot-dried). For Panavia F 2.0 (PF, Kuraray), the subgroups were: PF-control (uncontaminated), PF-I (saliva contamination, dried), PF-II (saliva contamination, rinse, dried), PF-III (primer, saliva contamination, dried), PF-IV (primer, saliva contamination, dried, primer re-applied) and PF-V (primer, saliva contamination, rinse, dried, primer re-applied). Composite blocks were luted onto dentin using the two cements. Bonded specimens were sectioned into 0.9 mm x 0.9 mm beams for muTBS testing. Representative fractured beams were prepared for fractographic analysis. RESULTS: For ARC, salivary contamination of etched dentin (ARC-I) significantly lowered bond strength (p=0.001). Rinsing saliva off with water (ARC-II) restored bond strength to control level. Re-etching dentin surface after rinsing (ARC-III) resulted in the lowest bond strength (p<0.001). For PF, salivary contamination of dentin before (PF-I) and after application of primer (PF-III and PF-IV) significantly lowered bond strength (p<0.001). Rinsing saliva off with water and re-application of primer (PF-II and PF-V) improved bond strength. CONCLUSION: Saliva contamination during luting deteriorated the bond quality of resin cements. Decontamination by rinsing with water was most effective in restoring the bond strength of RelyX ARC. Decontamination by water-rinsing and primer re-application after rinsing improved the bond strength of Panavia F 2.0.

High-level dosimetry at the demagnetization experiments of permanent magnets.

The measurements of high-energy and high dose mixed radiation from high-energy electron accelerator are carried out using a radiation damage monitor. It consists of two Radiation-Sensing Field-Effect Transistors (RADFETs) for total absorbed dose from mainly gamma ray and other charged particles and a Si PIN diode for neutron fluence. This is a part of the demagnetization study of rare earth permanent magnet irradiated by 2.5-GeV electron beam. The sensitivities of damage detectors are measured using 65-MeV quasi-monoenergic neutron, 14-MeV D-T neutron, (252)Cf neutron for Si PIN diode and (60)Co and (137)Cs gamma ray for RADFETs. Measured sensitivities are in acceptable range in the comparison of producer's proposed values. The dose and fluence measurements are carried out for the same target condition, Cu and Ta, as that for the demagnetization study. The 5 x 5 cm(2) cross-sectional and 5.5-cm-thick Pb target is also used for the general comparison with photoneutron yields. All measured dose and fluence are compared with the calculated results using the FLUKA code and agree well each other. The application of this kind of radiation damage monitor to high-level dosimetry at high-energy electron accelerator has been discussed.

Financial burden of national health insurance for treating patients with transfusion-dependent thalassemia in Taiwan.

The thalassemias are a heterogeneous group of inherited hypochromic anemias of varying severity. The mainstay of supportive treatment is regular blood transfusion accompanied by iron-chelating therapy. Hematopoietic stem cell transplantation (HSCT) provides an alternative option when curative therapy is considered. More than 400 patients in Taiwan have beta-thalassemia major or other transfusion-dependent thalassemias, and their treatment costs account for a considerable percentage of the National Health Insurance expenditure. In this report, we estimated the treatment costs of conventional therapy (regular blood transfusion accompanied by iron-chelating agents) and HSCT. The undiscounted medical cost of 20 years of follow-up (20 years from diagnosis) and the undiscounted total lifetime cost were NT$ 4 739 888 (NT$ means New Taiwan Dollars)/US$ 149 288 and NT$ 11 529 990/US$ 363 149, respectively, for patients undergoing conventional therapy, and NT$ 2 639 982/US$ 83 149 and NT$ 3 511 172/US$ 110 588, respectively, for those undergoing successful HSCT. Comparisons of treatment costs and other parameters between these two modalities can add to the information base on which policy is made by health authorities or clinicians.

Feasibility study of nitrogen removal with the mecellulose wasted liquor as an external carbon source in the two-stage denitrification process.

The utilization of mecellulose wasted liquor (MWL) as an external carbon source was investigated to find an alternative for methanol in the two-stage denitrification pilot process. The pilot plant was supplied with the raw water from the J-Municipal Sewage Treatment Plant (J-MSTP) in Korea. The raw water of J-Municipal Sewage Treatment Plant contains low and high concentration of biodegradable organics and nitrogen source, respectively, due to the inflow of industrial wastewater and landfill leachate. Methanol was fed to provide external carbon source for high concentration of nitrogen source removal by denitrification in this J-Municipal Sewage Treatment Plant, and thus this study was performed to test effects to the effluent quality and efficiencies of nitrogen source removal with an alternative carbon source for the cost reduction. The 6.5mg 1(-1) and 5.7mg l(-1) of total nitrogen (TN) concentration in the effluent were achieved with mecellulose and methanol, whereas mecellulose and methanol were fed to give the same ratio of gCODgNO,-N(-1), respectively. The 60% of COD in mecellulose wasted liquor was used as a carbon source for denitrification and the stable denitrification rate was earned when one half of the required total carbon source for denitrification was fed to pre-anoxic tank in the pilot plant. The required gCODgNO,-N(-1) ratio with mecellulose wasted liquor was 1.4 times higher than with methanol. Mecellulose wasted liquor is feasible to be used as external carbon source for organic loading, nitrogen and phosphorus removal. If mecellulose wasted liquor is considered as an alternative external carbon source to substitute methanol 26-28m3 mecellulose wasted liquor per 1 m3 methanol will be required. However, to meet with the effluent standard (10 mg BOD l(-1)) for J-Municipal Sewage Treatment Plant, the feed concentration of mecellulose wasted liquor should be recommended to be lower than 200 mgl(-1).

Increased mortality odds ratio of male liver cancer in a community contaminated by chlorinated hydrocarbons in groundwater.

AIMS: To investigate the association between cancer mortality risk and exposure to chlorinated hydrocarbons in groundwater of a downstream community near a contaminated site. METHODS: Death certificates inclusive for the years 1966-97 were collected from two villages in the vicinity of an electronics factory operated between 1970 and 1992. These two villages were classified into the downstream (exposed) village and the upstream (unexposed) according to groundwater flow direction. Exposure classification was validated by the contaminant levels in 49 residential wells measured with gas chromatography/mass spectrometry. Mortality odds ratios (MORs) for cancer were calculated with cardiovascular-cerebrovascular diseases as the reference diseases. Multiple logistic regressions were performed to estimate the effects of exposure and period after adjustment for age. RESULTS: Increased MORs were observed among males for all cancer, and liver cancer for the periods after 10 years of latency, namely, 1980-89, and 1990-97. Adjusted MOR for male liver cancer was 2.57 (95% confidence interval 1.21 to 5.46) with a significant linear trend for the period effect. CONCLUSION: The results suggest a link between exposure to chlorinated hydrocarbons and male liver cancer risk. However, the conclusion is limited by lack of individual information on groundwater exposure and potential confounding factors.

Use of SoftPerm contact lenses when rigid gas permeable lenses fail.

PURPOSE: We evaluated the performance of the SoftPerm contact lens (Wesley Jessen) in patients with irregular astigmatism, usually due to keratoconus or after penetrating keratoplasty (PK), who were unable to befitwith, or intolerant of, rigid gas permeable (RGP) contact lenses. METHODS: A retrospective study of patients fit with SoftPerm lenses in the Cornea Department at Wills Eye Hospital between March 1985 and March 2000 was performed. RESULTS: Thirty-five cases were reviewed, with follow-up available in 33 cases. Most of the eyes had irregular astigmatism secondary to keratoconus (22/35,62.9%) or PK (10/35,28.6%) and had failed a trial of RGP lenses. The mean logMAR visual acuity with SoftPerm lenses was 0.13+/-0.18 (range -0.12 to 0.6). In 25 cases in which comparison with glasses or RGP lenses was possible, SoftPerm lenses provided better visual acuity than glasses in 17/25 cases (68%) with a mean difference of -0.24 (P = 0.001, paired t-test); visual acuity with SoftPerm lenses was better than RGP visual acuity in 13/25 cases (52%), with a mean difference of -0.06 (P = 0.07, paired t-test). Complications included broken lenses (16/33,48.5%), giant papillary conjunctivitis (GPC) (9/33, 27.3%), and peripheral corneal neovascularization (9/33, 27.3%). The GPC and peripheral corneal neovascularization were often delayed in presentation. The major subjective complaint was discomfort (13/33, 39.4%). At the last follow-up, the SoftPerm lens was still in use in 22/33 cases (66.7%). Discomfort was the most common reason for discontinuation. The mean duration of lens wear was 52.5+/-31.7 months, range 3 to 110 months. CONCLUSIONS: The SoftPerm lens can provide satisfactory visual correction in many cases of irregular astigmatism with RGP failure. However, problems such as frequent breakage, GPC, peripheral corneal neovascularization, and discomfort necessitate close follow-up.

Comparison of balloon dacryocystoplasty to probing as the primary treatment of congenital nasolacrimal duct obstruction.

PURPOSE: In children older than 18 months, primary probing procedures for congenital nasolacrimal duct obstruction (CNLDO) are thought to have lower rates of success. This study compares the results of primary probing to balloon dacryocystoplasty (DCP) in children stratified by age category. METHODS: In a retrospective chart review, 29 eyes with CNLDO that underwent balloon DCP in children older than 18 months were identified and age-matched to 29 eyes that underwent probing. The eyes were divided into 3 age categories: category 1 (18-24 months), category 2 (24-36 months), and category 3 (>36 months). RESULTS: Of the 29 eyes treated with balloon DCP (mean age, 37.1 months), 26 were successfully treated. Twenty-five of the 29 matched probed eyes (mean age, 31.1 months) were successfully treated, resulting in an overall success rate of 90% for balloon DCP and 86% for primary probing. Within each age category, the success rate varied but did not show an advantage to balloon DCP. The presence of crusting and expressible discharge from the puncta during preoperative evaluation predicted a successful probing (OR, 16; 95% CI, 1.3-192). CONCLUSION: Overall, balloon DCP did not appear to present an advantage as compared with primary probing as the initial treatment in these children. Primary probing has an impressive overall success rate that did not diminish in the children older than 36 months.

Cleavage of Bax is mediated by caspase-dependent or -independent calpain activation in dopaminergic neuronal cells: protective role of Bcl-2.

Two cysteine protease families, caspase and calpain, are known to participate in cell death. We investigated whether a stress-specific protease activation pathway exists, and to what extent Bcl-2 plays a role in preventing drug-induced protease activity and cell death in a dopaminergic neuronal cell line, MN9D. Staurosporine (STS) induced caspase-dependent apoptosis while a dopaminergic neurotoxin, MPP(+) largely induced caspase-independent necrotic cell death as determined by morphological and biochemical criteria including cytochrome c release and fluorogenic caspase cleavage assay. At the late stage of both STS- and MPP(+)-induced cell death, Bax was cleaved into an 18-kDa fragment. This 18-kDa fragment appeared only in the mitochondria-enriched heavy membrane fraction of STS-treated cells, whereas it was detected exclusively in the cytosolic fraction of MPP(+)-treated cells. This proteolytic cleavage of Bax appeared to be mediated by calpain as determined by incubation with [(35)S]methionine-labelled Bax. Thus, cotreatment of cells with calpain inhibitor blocked both MPP(+)- and STS-induced Bax cleavage. Intriguingly, overexpression of baculovirus-derived inhibiting protein of caspase, p35 or cotreatment of cells with caspase inhibitor blocked STS- but not MPP(+)-induced Bax cleavage. This appears to indicate that calpain activation may be either dependent or independent of caspase activation within the same cells. However, cotreatment with calpain inhibitor rescued cells from MPP(+)-induced but not from STS-induced neuronal cell death. In these paradigms of dopaminergic cell death, overexpression of Bcl-2 prevented both STS- and MPP(+)-induced cell death and its associated cleavage of Bax. Thus, our results suggest that Bcl-2 may play a protective role by primarily blocking drug-induced caspase or calpain activity in dopaminergic neuronal cells.

FLASH coordinates NF-kappa B activity via TRAF2.

FLASH is a protein recently shown to interact with the death effector domain of caspase-8 and is likely to be a component of the death-inducing signaling complex in receptor-mediated apoptosis. Here we show that antisense oligonucleotide-induced inhibition of FLASH expression abolished TNF-alpha-induced activation of NF-kappaB in HEK293 cells, as determined by luciferase reporter gene expression driven by a NF-kappaB responsive promoter. Conversely, overexpression of FLASH dose-dependently activated NF-kappaB, an effect suppressed by dominant negative mutants of TRAF2, NIK, and IKKalpha, and partially by those of TRAF5 and TRAF6. TRAF2 was co-immunoprecipitated with FLASH from the cell extracts of HEK293 cells or HeLa cells stably expressing exogenous FLASH (HeLa/HA-FLASH). Furthermore, serial deletion mapping demonstrated that a domain spanning the residues 856-1191 of FLASH activated NF-kappaB as efficiently as the full-length and could directly bind to TRAF2 in vitro and in the transfected cells. Taken together, these results suggest that FLASH coordinates downstream NF-kappaB activity via a TRAF2-dependent pathway in the TNF-alpha signaling.

Human immunodeficiency virus p24 antigen testing in cornea donors.

PURPOSE: Testing for the p24 antigen of the human immunodeficiency virus (HIV) may detect early HIV infection in the seronegative window; however, falsely reactive results may occur in cadaver specimens. Although neither the Food and Drug Administration (FDA) nor the Eye Bank Association of America requires p24 testing of cornea donors, many tissue banks using other organs from cornea donors do perform this assay, and the FDA requires that eye banks reject corneal tissue if a reactive p24 assay is reported. We investigated the impact of p24 testing on eye banking and corneal transplantation. METHODS: Two clinical cases and records from the Lions Eye Bank of Delaware Valley (LEBDV) were reviewed retrospectively. RESULTS: Two corneas from the LEBDV were transplanted before the reporting of p24 reactivity by other tissue banks. In one case, because of the young age of the recipient, the surgeon elected to replace the cornea with new tissue hours after the original transplant, and later polymerase chain reaction (PCR) testing was negative. In the other case, there was not enough specimen to perform Western blot or PCR confirmatory testing. The patient was followed with periodic serologic testing for HIV and has remained seronegative. To avoid such problems in the future, the LEBDV initiated testing of all donors with p24 and other nonrequired screening tests. Over a 2-month period, 22 corneas (from 11 donors) were discarded because of these tests: 4 donors had reactive p24 tests, 6 were reactive for antibody to hepatitis B core antigen, and 1 had a reactive syphilis test. CONCLUSIONS: Results from p24 assays by other tissue banks may cause difficult clinical situations when the results are received after transplantation of the tissue, but the use of the p24 assay in the screening of cornea donors may result in excessive waste of donor tissue. Further guidance is needed regarding the management of positive results from this and other nonrequired screening tests.

Inactivation of farnesyltransferase and geranylgeranyltransferase I by caspase-3: cleavage of the common alpha subunit during apoptosis.

Caspase plays an important role in apoptosis. We report here that farnesyltransferase/geranylgeranyltransferase (FTase/GGTase)-alpha, a common subunit of FTase (alpha/beta(FTase)) and GGTase I (alpha/beta(GGTase)), was cleaved by caspase-3 during apoptosis. FTase/GGTase-alpha (49 kDa) was cleaved to 35 kDa (p35) in the Rat-2/H-ras, W4 and Rat-1 cells treated with FTase inhibitor (LB42708), anti-Fas antibody and etoposide, respectively. This cleavage was inhibited by caspase-inhibitors (YVAD-cmk, DEVD-cho). Serial N-terminal deletions and site-directed mutagenesis showed that Asp59 of FTase/GGTase-alpha was cleaved by caspase-3. The common FTase/GGTase-alpha subunit, but not the beta subunits, of the FTase or GGTase I protein complexes purified from baculovirus-infected SF-9 cells was cleaved to be inactivated by purified caspase-3. In contrast, FTase mutant protein complex [(D(59)A)alpha/beta(FTase)] was resistant to caspase-3. Expression of either the cleavage product (60-379) or anti-sense of FTase/GGTase-alpha induced cell death in Rat-2/H-ras cells. Furthermore, expression of (D(59)A)FTase/GGTase-alpha mutant significantly desensitized cells to etoposide-induced death. Taken together, we suggest that cleavage of prenyltransferase by caspase contributes to the progression of apoptosis.

Penetrating keratoplasty in iridocorneal endothelial syndrome.

PURPOSE: To evaluate the clinical outcome of penetrating keratoplasty (PK) in iridocorneal endothelial (ICE) syndrome. METHODS: Clinical charts of patients who underwent penetrating keratoplasty for ICE syndrome between 1985 and 1999 were reviewed retrospectively. Glaucoma control, best corrected visual acuity pre- and post-PK, graft clarity, graft rejection episodes, improvement in pain, and additional procedures were analyzed. RESULTS: Fourteen cases were reviewed with an average follow-up of 58 months after PK. Initial grafts failed in seven patients (50%), in six cases because of rejection, and one owing to endothelial failure without signs of rejection. Repeat PKs were performed in six patients. At final follow-up, 12 grafts were clear. Glaucoma was controlled pre- and post-PK (average intraocular pressure, 16 mmHg for both eyes). Pre-PK, eight patients were using glaucoma medicines and nine had had glaucoma surgery. At the end of the follow-up, seven patients were using glaucoma medicines; six patients required glaucoma surgery after their initial PK. At the final follow-up visit, visual acuity in three patients (21%) was 20/40 or better, it ranged from 20/50 to 20/100 in four patients (29%) and 20/200 to 20/400 in five patients (36%), and in two patients with failed grafts (14%) it was counting fingers or worse. CONCLUSION: Clear grafts were achieved in 12 cases, although six patients (43%) underwent repeat PKs. All patients had glaucoma, which was controlled before and after PK by medical treatment and surgical procedures. Favorable outcomes can be achieved in patients with ICE syndrome but may require multiple corneal and glaucoma procedures.

Pro-apoptotic function of calsenilin/DREAM/KChIP3.

Apoptotic cell death and increased production of amyloid b peptide (Ab) are pathological features of Alzheimer's disease (AD), although the exact contribution of apoptosis to the pathogenesis of the disease remains unclear. Here we describe a novel pro-apoptotic function of calsenilin/DREAM/KChIP3. By antisense oligonucleotide-induced inhibition of calsenilin/DREAM/KChIP3 synthesis, apoptosis induced by Fas, Ca2+-ionophore, or thapsigargin is attenuated. Conversely, calsenilin/DREAM/KChIP3 expression induced the morphological and biochemical features of apoptosis, including cell shrinkage, DNA laddering, and caspase activation. Calsenilin/DREAM/KChIP3-induced apoptosis was suppressed by caspase inhibitor Z-VAD and by Bcl-XL, and was potentiated by increasing cytosolic Ca2+, expression of Swedish amyloid precursor protein mutant (APPSW) or presenilin 2 (PS2), but not by a PS2 deletion lacking its C-terminus (PS2/411stop). In addition, calsenilin/DREAM/KChIP3 expression increased Ab42 production in cells expressing APPsw, which was potentiated by PS2, but not by PS2/411stop, which suggests a role for apoptosis-associated Ab42 production of calsenilin/DREAM/KChIP3.

An anti-apoptotic protein human survivin is a direct inhibitor of caspase-3 and -7.

Survivin, an apoptosis inhibitor/cell-cycle regulator, is critically required for suppression of apoptosis and ensuring normal cell division in the G2/M phase of the cell cycle. It is highly expressed in a cell cycle-regulated manner and localizes together with caspase-3 on microtubules within centrosomes. Whether survivin is a physiologically relevant caspase inhibitor has been unclear due to the difficulties with obtaining correctly folded survivin and finding the right conditions for inhibition assay. In this study, recombinant, active human survivin was expressed in Escherichia coli and purified to homogeneity. The protein, existing as a homodimer in solution, binds caspase-3 and -7 tightly with dissociation constants of 20.9 and 11.5 nM, respectively, when evaluated by surface plasmon resonance spectroscopy. Consistently, survivin potently inhibits the cleavage of a physiological substrate poly(ADP-ribose) polymerase and an artificial tetrapeptide by caspase-3 and -7 in vitro with apparent inhibition constants of 36.0 and 16.5 nM, respectively. The data suggest that sequestering caspase-3 and -7 in inhibited states on microtubules is at least one mechanism of survivin in the suppression of default apoptosis in the G2/M phase. The localization of survivin on microtubules, which is essential for its function, should increase the protective activity at the action site.

Proapoptotic effects of tau cleavage product generated by caspase-3.

Using an in vitro translation assay to screen a human brain cDNA library, we isolated the microtubule-associated protein Tau and determined it to be a caspase-3 substrate whose C-terminal cleavage occurred during neuronal apoptosis. DeltaTau, the 50-kDa cleavage product, was detected by Western blot in apoptotic cortical cells probed with anti-PHF-1 and anti-Tau-5 antibodies, but not anti-T-46 antibody which recognizes the C-terminus. Overexpression of DeltaTau in SK-N-BE2(C) cells significantly increased the incidence of cell death. Staurosporine-induced Tau cleavage was blocked by 20 microM z-Asp-Glu-Val-Asp-chloromethylketone, a caspase-3 inhibitor, and in vitro, Tau was selectively cleaved by caspase-3 or calpain, a calcium-activated protease, but not by caspases-1, -8, or -9. (D421E)-Tau, a mutant in which Asp421 was replaced with a Glu, was resistant to cleavage by caspase-3 and tended to suppress staurosporine-induced cell death more efficiently than did wild-type Tau in both transient and stable expression systems. Finally, the incidence of DeltaTau-induced cell death was augmented by expression of Abeta precursor protein (APP) or Swedish APP mutant. Taken together, these results suggest that the caspase-3 cleavage product of Tau may contribute to the progression of neuronal cell death in Alzheimer's disease.

Cometabolic biosynthesis of copolyesters consisting of 3-hydroxyvalerate and medium-chain-length 3-hydroxyalkanoates by Pseudomonas sp. DSY-82.

A newly isolated strain, designated as Pseudomonas sp. DSY-82, synthesized medium-chain-length polyhydroxyalkanoate (MCL-PHA) copolyesters when grown on alkanoates from hexanoate to undecanoate as the sole carbon source. When used alone, butyrate and valerate supported the growth of the isolate but not PHA production. However, unusual polyesters containing 3-hydroxyvalerate, as well as various MCL 3-hydroxyalkanoate monomeric units, were synthesized when valerate was cofed with either nonanoate or 10-undecenoate, suggesting the formation of monomer units from both substrates. Concentrations of 3-hydroxyvalerate, 3-hydroxyoctanoate, and 3-hydroxydecanoate in the PHAs produced were significantly elevated by the addition of valerate, indicating that the incorporation of these monomer units to PHA occurred primarily through cometabolism. The total amount of these monomer units in the PHAs reached up to 30%. The PHAs produced in this study were most likely random copolyesters as determined by differential scanning calorimetric analysis. This is the first case of microbial synthesis of copolyesters consisting of 3-hydroxyvalerate and MCL 3-hydroxyalkanoate monomer units through cometabolism.

Reconstitution of caspase-8 sensitizes JB6 cells to TRAIL.

TRAIL induces apoptosis in various tumor cells. We report here that caspase-8 is required in TRAIL-induced cell death. Western blot analyses and enzyme assays showed that exposing Jurkat cells to TRAIL resulted in activation of caspases-8 followed by caspase-3 and -9. Acetyl-IETD-fluoromethylketone, a caspase-8 inhibitor, potently suppressed TRAIL-induced cell death compared to acetyl-DEVD-fluoromethylketone and acetyl-LEHD-fluoromethylketone, inhibitors of caspase-3 and caspase-9, respectively. JB6 cells, a caspase-8-deficient Jurkat variant, were completely resistant to TRAIL. However, reconstitution with a caspase-8, but not with caspase-2 or -3, sensitized JB6 cells to subsequent exposure to TRAIL. These results are indicative of the crucial function of caspase-8 in TRAIL-induced apoptosis in Jurkat cells.