Serum creatine kinase activity following forearm flexion isometric exercise

Summary The purpose of this study was to 1) compare serum creatine kinase (CK) activity following two forearm flexion isometric exercise regimens differing in work to rest ratio, and 2) examine the CK response to a repeated bout of isometric exercise. Eleven males were tested on two sessions (bouts) spaced 1 week apart. For bout 1, five subjects (group A) performed a forearm flexion isometric exercise consisting of 40 10-s maximal contractions with 20-s inter-trial rests (1020), while six (group B) performed 40 maximal 10-s contractions with 5-s inter-trial rests (105). The increase in serum CK activity following the 1020 exercise (143%) was significantly greater than that following the 105 exercise (52%). The 1020 exercise was also associated with greater tension generation over trials. One week later, both groups performed a bout of 1020 exercise. A substantial reduction in the serum CK response was found following this second bout. The data suggest that for bout 1 the isometric exercise associated with the greater overall tension levels resulted in the greater CK response. However, when the 1020 exercise was repeated 1 week later, a substantial reduction in the CK response was found which was unrelated to the tension generated.This study was supported by a University Faculty Research Grant No. 2-03021     

Site-specific mutagenicity of stereochemically defined 1,N2-deoxyguanosine adducts of trans-4-hydroxynonenal in mammalian cells

Trans-4-hydroxynonenal (HNE) is a toxic compound produced endogenously during lipid peroxidation. HNE is a potent electrophile that is reactive with both proteins and nucleic acids. HNE preferentially reacts with deoxyguanosine to form four stereoisomeric HNE-deoxyguanosine (HNE-dG) adducts: (6R, 8S, 11R), (6S, 8R, 11S), (6R, 8S, 11S), and (6S, 8R, 11R). These adducts were synthesized into 12-mer oligodeoxynucleotides, inserted into a DNA shuttle vector and evaluated for the ability of each stereoisomer to induce mutagenesis when replicated through mammalian cells. The resultant mutagenicity of these adducts was related to their stereochemistry, in that two of the HNE-dG adducts, (6R, 8S, 11R) and (6S, 8R, 11S), were significantly more mutagenic than the (6R, 8S, 11S) and (6S, 8R, 11R) HNE-dG adducts. These data conclusively demonstrate that HNE-derived DNA adducts can be mutagenic in mammalian cells and their ability to cause mutations is dictated by their stereochemistry.     

Antitumor efficacy of regional oncolytic viral therapy for peritoneally disseminated cancer

Oncolytic viral therapy is a promising new method of cancer treatment. Peritoneal dissemination of cancer is a common and fatal clinical condition seen in many malignancies, with few effective therapies available. G207, a multimutated replication-competent herpes simplex virus type-1, effectively treats disseminated peritoneal cancer. This study evaluates viral proliferation and subsequent tumoricidal effects in vitro and in vivo after regional viral delivery. In vitro studies demonstrate that G207 efficiently kills five human gastric cancer cell lines, and that permissiveness to viral replication is correlated with cytotoxicity. In a murine xenograft model of human gastric carcinomatosis, peritoneal delivery of G207 effectively kills tumor and prolongs survival. Data from quantitative PCR characterizes peritoneal clearance of virus after intraperitoneal injection, and identifies G207 replication within tumor cells in vivo, similar to in vitro proliferation. Further analysis of various organs confirms that G207 does not replicate within normal tissue after peritoneal delivery. Wild-type KOS viral replication was also demonstrated in vivo, with significant toxicity secondary to dissemination and encephalitis. In vivo viral proliferation of G207 is restricted to tumor cells, is correlated with in vitro assays, and is an important mechanism of anticancer efficacy.     

Synthesis and Characterization of Segmented Poly(ether ester)s Containing H‐Bonding Units

A series of poly(ether ester)s containing different H‐bonding units (amide, carbamate, urea) was prepared by polycondensation in bulk, using Ti(OBu)₄ as a catalyst. The copolymers were obtained starting from PEG1000, 1,4‐butanediol, and a symmetrical, bis‐ester terminated monomer carrying H‐bonding units. These materials were designed for biomedical applications, in which ultimate biodegradability of the materials is required. The influence of the nature of the H‐bonding unit and of the length of the methylene spacer between H‐bonding groups on the thermal and solubility properties of copolymers was investigated. Amide containing copolymers were more thermally stable than ones containing carbamate, consistent with the observed behavior of the corresponding monomers. In most cases, differential scanning calorimetry (DSC) traces were quite complex because of phase separation and dependent on the applied cooling rate. Copolymers containing urea bonds were less soluble in most organic solvents, but their thermal properties were not significantly different than their amide containing counterparts.

Synthesis of amide‐based diester monomers.     

Effect of dimethyl sulfoxide on the in vitro and in vivo bactericidal activity of human and mouse neutrophils and mononuclear phagocytes

Dimethyl sulfoxide (DMSO) is reported to have antiinflammatory activity in various systems. Since resistance to bacterial infection can be thought of as a specialized type of inflamation, we were interested in determining the effect of DMSO on phagocyte bactericidal activity. The results indicated that in vitro DMSO treatment of human and mouse neutrophils and mononuclear phagocytes caused a dose-dependent inhibition of the killing ofEscherichia coli andListeria monocytogenes. However, pretreatment of mice with DMSO in vivo caused only a slight decrease in the subsequent in vitro bactericidal activity of neutrophils and macrophages from those mice. In addition, repeated injection of mice with a physiologically relevant dosage of DMSO did not enhance the lethality of eitherE. coli orL. monocytogenes, nor did it affect the clearance of a sublethalListeria challenge from the spleen and liver. These results suggest that clinical usage of DMSO should not predispose human subjects to bacterial infection.This work was supported by National Institutes of Health grants AI-11240 and GM-24834.     

Anti-SARS-CoV-2 activity of targeted kinase inhibitors: Repurposing clinically available drugs for COVID-19 therapy

Coronavirus disease 2019 (COVID-19) remains a major public health concern, and vaccine unavailability, hesitancy, or failure underscore the need for discovery of efficacious antiviral drug therapies. Numerous approved drugs target protein kinases associated with viral life cycle and symptoms of infection. Repurposing of kinase inhibitors is appealing as they have been vetted for safety and are more accessible for COVID-19 treatment. However, an understanding of drug mechanism is needed to improve our understanding of the factors involved in pathogenesis. We tested the in vitro activity of three kinase inhibitors against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), including inhibitors of AXL kinase, a host cell factor that contributes to successful SARS-CoV-2 infection. Using multiple cell-based assays and approaches, gilteritinib, nintedanib, and imatinib were thoroughly evaluated for activity against SARS-CoV-2 variants. Each drug exhibited antiviral activity, but with stark differences in potency, suggesting differences in host dependency for kinase targets. Importantly, for gilteritinib, the amount of compound needed to achieve 90% infection inhibition, at least in part involving blockade of spike protein-mediated viral entry and at concentrations not inducing phospholipidosis (PLD), approached a clinically achievable concentration. Knockout of AXL, a target of gilteritinib and nintedanib, impaired SARS-CoV-2 variant infectivity, supporting a role for AXL in SARS-CoV-2 infection and supporting further investigation of drug-mediated AXL inhibition as a COVID-19 treatment. This study supports further evaluation of AXL-targeting kinase inhibitors as potential antiviral agents and treatments for COVID-19. Additional mechanistic studies are needed to determine underlying differences in virus response.     

Dose-Dependent Carcinogenicity and Frequent Ki-ras Proto-oncogene Activation by Dietary N-Nitrosodiethylamine in Rainbow Trout

While the experimental data upon which current concepts in mechanisticallybased risk assessment and molecular epidemiology are groundedderive almost entirely from rodent models, fish models haveseveral attributes (e.g., low background incidence, extremelylow cost tumor studies, nonmammalian comparative status forextrapolation of mechanisms to humans) that make them valuableadjuncts for addressing these concepts. This report providesan initial characterization of the dose dependency of dietaryN-nitrosodiethylamine (DEN) hepatocarcinogenicity in Shastastrain rainbow trout (Oncorhynchus mykiss) and the potentialof DEN to elicit ras proto-oncogene activation in this species.Carcinogen was administered in the diet at five concentrationsfor 12 months. Necropsies were per formed at 9, 12, and 18 months,the latter on fish maintained on control diet for 6 months aftercessation of DEN exposure. The incidence of hepatic neoplasmsat the lower dietary concentrations (70 ppm) did not consistentlyexceed that for control groups, which were higher in this particularstudy (2%) than expected (historically 0.1%). For the higherDEN concentrations, a linear relationship between the hepatictumor incidence (expressed as log odds, log [p/(1-p)1, wherep = proportion of fish bearing tumors), and the logarithm oftotal cumulative dose was observed, with response being independentof the length of time (9 or 12 months) during which the dosewas accumulated. The dose-response curve for fish maintainedan additional 6 months postexposure was shifted toward higherincidence but was parallel to the curve for fish killed at cessationof exposure. The model predicts that doubling the dose willproduce some what more than a doubling of the odds (pl(100 -p) for tumor incidence and that the odds for lesions 6 monthspostexposure will be approximately double those at cessationof exposure. Comparison of these results with previous studiesusing rats suggests an overall similarity in dose-response curves,with trout being somewhat less sensitive than rats to DEN hepatocarcinogenesis. To examine the molecular basis for DEN carcinogenesis in this species, seven liver tumors induced separatelyby short-term DEN treatment were probed by 3'-mismatch primerpolymerase chain reaction analysis for evidence of Ki-ras proto-oncogeneactivating point mutations. A very high proportion (6/7) oftumors was found to carry codon 12 GGA - AGA mutations, whereasno codon 61 mutants were detected in this sample. These initialresults differ from those reported using hepatic tumors fromDEN-treated mice, which exhibit frequent Ha-ras codon 61 mutations[Richardson et al., Carcinogenesis 13, 1277–1279 (1992)]and rats, which appear not to carry DEN-activated ras alleles[Bauer-Hoffman et al., Carcinogenesis 11, 1875–1877 (1990)].Thus the available oncogene data for the common carcinogen DENdo not suggest a simple, consistent oncogenic pathway or mutationalspectrum useful in the molecular epidemiology of human cancers.     

Linking Clients from HIV Antibody Counseling and Testing to Prevention Services

The effectiveness of HIV antibody counseling and testing as a prevention intervention is limited: persons testing seronegative do not usually change their risk behaviors, some actually increase their risk behaviors, and decreases in risk behaviors are usually short-lived. Referrals to additional prevention and other needed services are therefore recommended, although the extent and determinants of referral provision for persons testing seronegative are unknown. We assessed the prevalence of referrals and the association between risk behaviors and prevention referrals among seronegatives. We reviewed HIV testing and referral data on all persons receiving confidential seronegative test results in San Francisco (SF) in the first 10 months of 1995 (n = 5,595), and gathered more detailed referral information at the municipal STD clinic from November 1995 through May 1996 (n = 747). The overall prevalence of referrals was low: a referral was given to 19.1% of the SF sample and 10.6% of the STD clinic sample; 15.4% of the SF sample and 5.9% of the STD clinic sample received a prevention referral. Injection drug users (IDUs) were the most likely to receive a prevention referral (48.5% of SF IDUs, 36.4% of STD clinic IDUs); men having sex with men and women with high-risk partners were also more likely to get a prevention referral than others. For SF IDUs, unsafe sex and needle sharing were not associated with an increased likelihood of receiving a prevention referral. Opportunities to link high-risk clients from counseling and testing to HIV prevention services are being missed. The referral component of HIV counseling and testing should be improved.     

Stereopsis and long-term stability of alignment in esotropia.

PURPOSE: Recent studies of infantile and accommodative esotropia (ET) have focused on stereoacuity as a final outcome measurement for judging the success or failure of treatment. The purpose of the present study was to extend this approach by evaluating whether the presence of stereopsis developing immediately after surgical alignment or optical correction plays a role in maintenance of long-term alignment. METHODS: Random-dot stereoacuity was assessed within 3 months of initial surgical alignment in 70 children with infantile ET and within 3 months of initial optical correction in 66 children with accommodative ET. At > or = 5 years of age, adverse outcomes were assessed including loss of alignment, amblyopia, and nil stereopsis. Risk-factor analysis was used to evaluate whether early nil stereopsis increased the risk for subsequent adverse outcomes. RESULTS: In the infantile ET cohort, early nil stereopsis was associated with a 3.6 times (95% confidence interval [CI] 2.4 to 4.1) greater risk of surgery for recurrent ET or consecutive exotropia and a 4.2 times (95% CI 3.3 to 4.4) greater risk for nil stereopsis at > or = 5 years of age. In the accommodative ET cohort, early nil stereopsis was associated with a 17.4 times (95% CI 3.3 to 32.2) greater risk of surgery for ET and a 32.2 times (95% CI 15.8 to 35.6) greater risk for nil stereopsis at > or = 5 years of age. CONCLUSION: Treatment protocols designed to optimize stereoacuity outcomes promote long-term stability of alignment.