Patient's understanding of and satisfaction with acute stroke treatment.

OBJECTIVE: We aimed to assess patient and surrogate understanding of and satisfaction with communication regarding acute stroke treatments of intravenous thrombolysis (IVT) and endovascular therapy (EVT). METHODS: In this single health-system prospective observational study, patients or their surrogates were interviewed within 72 h of acute stroke therapy. Respondent's satisfaction and self-reported understanding were rated on a Likert scale. Responses to open-ended questions were evaluated for recall of purpose and risks of treatment. RESULTS: Of 56 completed interviews (24 patients and 32 surrogates), 33 patients received IVT alone, 11 IVT and EVT, 12 EVT alone. Forty participants (71%) reported being extremely satisfied with their acute stroke care, 46 (82%) reported no difficulty understanding the purpose of treatment, while 36 (64%) reported no difficulty understanding risks. Two or more risks were verbalized by 8 (24%) participants for IVT, 2 (17%) for EVT, and 7 (64%) for both IVT and EVT. Brain bleeding was the most recalled risk for IVT and "lack of benefit" for EVT. CONCLUSIONS: Majority of the participants were extremely satisfied and reported no difficulty understanding purpose and risks of acute stroke treatment, however only 30% were able to verbalize two or more risks associated with the treatment.

The impact of COVID-19 on otolaryngology research: a cross-sectional analysis of discontinued trials.

CONTEXT: The COVID-19 pandemic has reduced the capacity to conduct medical research due to recruitment difficulties, supply chain shortages, and funding deficits. The clinical practice of otolaryngology was especially impacted due to a reduction in elective procedures, such as facial plastic surgeries and vocal fold injections. OBJECTIVES: The primary objective was to examine the extent of clinical trial (CTs) disruption secondary to the COVID-19 pandemic in the field of otolaryngology. METHODS: On August 1, 2021, we conducted a systematic search utilizing ClinicalTrials.gov for CTs related to common otolaryngology disorders. We utilized the date range January 1, 2020 through August 1, 2021 to identify all trials potentially affected by the COVID-19 pandemic. Investigators performed screening and data extraction in a duplicate, masked fashion. Trials resulting from the search were extracted for trial status, condition treated, enrollment number, funding, study type, study design, last update posted date, and trial location. Trials that explicitly mentioned COVID-19 as a reason for discontinuation or suspension were coded as such. For trials that did not explicitly mention COVID-19, we coded the reason provided from ClinicalTrials.gov. The Oklahoma State University Center for Health Science Institutional Review Board determined that this project did not qualify as human subject research. RESULTS: A total of 1,777 CTs met the inclusion criteria, and 223 CTs were discontinued between January 1, 2020 and August 1, 2021. Thirty-three (14.8%) of the 223 CTs reported discontinuation explicitly due to the COVID-19 pandemic. The 33 studies had 1,715 participants enrolled in total. Among the primary interventions, 11 (33.3%) were devices, 10 (30.3%) were drugs, 5 (15.2%) were behavioral, 4 (12.1%) were diagnostic tests, 1 (3.0%) was dietary, and 2 (6.1%) were labeled as "other." Regarding the CT location, 20 (60.6%) were conducted in the United States, and 13 (39.4%) were conducted internationally. Of the 33 CTs, 19 (57.6%) were suspended, 9 (27.3%) were terminated, and 5 (15.2%) were withdrawn. The overall most common reason for trial disruption was recruitment difficulties (24.2%). Median enrollment for discontinued trials due to COVID-19 was 37 (interquartile range [IQR], 19-71) and for other reasons was 6 (IQR, 0-27), for which the Mann-Whitney test showed a statistically significant difference between the two (z=-3.913, p<0.001). There were no significant associations between trial location, funding source, randomization, or whether a study involved masked vs unmasked participants. CONCLUSIONS: The COVID-19 pandemic has incited an impact on clinical research in the field of otolaryngology. To preserve trial continuation amid future threats to participant interaction and communication, we recommend further exploration of remote monitoring practices and virtual procedures-those that will maintain the effectiveness and accuracy needed to establish novel therapeutics. We encourage future trials to gauge which remote assessments show the greatest validity, with the long-term goal of establishing innovative study designs resilient to future pandemics.

Endemic North American Plants as Potentially Suitable Agents for Wound Cleaning Under Resource Scarce Conditions.

INTRODUCTION: Skin and subcutaneous infections are dangerous sequelae of soft tissue injuries, especially in austere situations where medical technology is not available. Numerous plant species endemic to North America have been described as having antibacterial properties. Of these, St. John's wort (Hypericum perforatum), chamomile (Matricaria chamomilla), and white oak (Quercus alba) were selected for testing against Staphylococcus aureus. Our objective was to assess the suitability of all 3 plants as potential antiseptic agents using methods easily replicated in a resource-scarce environment. METHODS: Water-soluble natural products were extracted from different concentrations of each plant part using either mechanical agitation at ambient temperature or boiling in unsterilized tap water. Antibacterial activity of each extract against S aureus was assessed using a conventional agar well diffusion bioassay. Zones of inhibition were measured using electronic calipers and were compared to tap water as the negative control. RESULTS: Aqueous extracts of St. John's wort and white oak bark displayed antibacterial effects against S aureus, with St. John's wort being more potent. Chamomile displayed no inhibitory properties at the concentrations examined. CONCLUSIONS: These data suggest that both St. John's wort and white oak are potential candidates for infection prophylaxis and therapy in austere wilderness scenarios, with St. John's wort being the more potent agent. White oak may be more logistically feasible because the larger surface area of a white oak tree allows for harvesting a larger quantity of bark compared to the smaller surface area of the St. John's wort plant.

The relationship between nicotine metabolism and nicotine and carcinogen exposure among American Indian commercial cigarette smokers and electronic nicotine delivery system users.

BACKGROUND: In American Indian (AI) tobacco users from the southern plains region of the US, we examined the relationship between nicotine and carcinogen exposure and nicotine metabolism. METHODS: Smokers (n = 27), electronic nicotine delivery system (ENDS) users (n = 21), and dual users (n = 25) of AI descent were recruited from a southern plains state. Urinary biomarkers of nicotine metabolism (nicotine metabolite ratio [NMR]), nicotine dose (total nicotine equivalents [TNE]), and a tobacco-specific lung carcinogen (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol and its glucuronides [total NNAL] were measured. RESULTS: The geometric mean of NMR was 3.35 (95% Confidence Interval(CI): 2.42, 4.65), 4.67 (95% CI: 3.39, 6.43), and 3.26 (95% CI: 2.44, 4.37) among smokers, ENDS users, and dual users. Each of the three user groups had relatively low levels of TNE, indicative of light tobacco use. Among smokers, there were inverse relationships between NMR and TNE (r = -0.45) and between NMR and NNAL (r = -0.50). Among dual users, NMR and TNE, and NMR and NNAL were not associated. Among ENDS users, NMR and TNE were not associated. CONCLUSIONS: AI tobacco users with higher NMR did not have higher TNE or NNAL exposure than those with lower NMR. This supports prior work among light tobacco users who do not alter their tobacco consumption to account for nicotine metabolism. IMPACT: The high prevalences of smoking and ENDS among AI in the southern plains may not be related to nicotine metabolism. Environmental and social cues may play a more important role in light tobacco users and this may be particularly true among AI light tobacco users who have strong cultural ties.

Biomarkers of Exposure in ENDS Users, Smokers, and Dual Users of American Indian Descent.

Objectives: We measured biomarkers of exposure among American Indian (AI) ENDS users, smokers, and dual users. Methods: Urine was analyzed for total nicotine equivalents (TNE) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol(NNAL). Expired-air carbon monoxide (CO) was collected. Two analyses were performed. "CO analysis" included smokers and dual users whose CO was ≥ 6 ppm and ENDS users whose CO was < 6 ppm. "NNAL analysis" included smokers and dual users whose NNAL was ≥ 47.3 pg/mg, and ENDS users whose NNAL was < 47.3 pg/mg. Biomarkers were summarized by geometric means (GM) and compared with nonparametric tests. Results: In both analyses, TNE was no different across the groups, and NNAL and CO were lower in ENDS users. In the NNAL analysis the GM of NNAL was 261.4, 6.1, and 228.0 pg/mg among smokers, ENDS users, and dual users (p < .001). Also in the NNAL analysis, the GM of CO was 14.7, 2.4, and 16.8 ppm among smokers, ENDS users, and dual users (p < .001). Conclusions: ENDS users did not differ in nicotine and had lower exposure to a lung carcinogen and a cardiovascular toxicant than smokers or dual users. Dual users and smokers did not differ in biomarker levels. Results should be used to inform tribal regulations and to educate the AI community on ENDS.

Community-Based Study Recruitment of American Indian Cigarette Smokers and Electronic Cigarette Users.

Data on the effectiveness of strategies for the recruitment of American Indians (AIs) into research is needed. This study describes and compares methods for identifying and recruiting AI tobacco users into a pilot study. Community-based strategies were used to recruit smokers (n = 35), e-cigarette users (n = 28), and dual users (n = 32) of AI descent. Recruitment was considered proactive if study staff contacted the individual at a pow wow, health fair, or vape shop and participation on-site or reactive if the individual contacted the study staff and participation occurred later. Screened, eligible, participated and costs and time spent were compared with Chi square tests. To understand AI descent, the relationship between number of AI grandparents and AI blood quantum was examined. Number of participants screened via the proactive strategy was similar to the reactive strategy (n = 84 vs. n = 82; p-value = 0.8766). A significantly greater proportion of individuals screened via the proactive than the reactive strategy were eligible (77 vs. 50%; p-value = 0.0002) and participated (75 vs. 39%; p-value = < 0.0001). Per participant cost and time estimated for the proactive strategy was $89 and 87 min compared to $79 and 56 min for the reactive strategy. Proportion at least half AI blood quantum was 32, 33, and 70% among those with 2, 3, and 4 AI grandparents, respectively (p = 0.0017). Proactive strategies resulted in two-thirds of the sample, but required more resources than reactive strategies. Overall, we found both strategies were feasible and resulted in the ability to reach sample goals. Lastly, number of AI biological grandparents may be a good, non-invasive indicator of AI blood quantum.

Multiyear Survey of Coccidia, Cryptosporidia, Microsporidia, Histomona, and Hematozoa in Wild Quail in the Rolling Plains Ecoregion of Texas and Oklahoma, USA.

We developed nested PCR protocols and performed a multiyear survey on the prevalence of several protozoan parasites in wild northern bobwhite (Colinus virginianus) and scaled quail (Callipepla squamata) in the Rolling Plains ecoregion of Texas and Oklahoma (i.e. fecal pellets, bird intestines and blood smears collected between 2010 and 2013). Coccidia, cryptosporidia, and microsporidia were detected in 46.2%, 11.7%, and 44.0% of the samples (n = 687), whereas histomona and hematozoa were undetected. Coccidia consisted of one major and two minor Eimeria species. Cryptosporidia were represented by a major unknown Cryptosporidium species and Cryptosporidium baileyi. Detected microsporidia species were highly diverse, in which only 11% were native avian parasites including Encephalitozoon hellem and Encephalitozoon cuniculi, whereas 33% were closely related to species from insects (e.g. Antonospora, Liebermannia, and Sporanauta). This survey suggests that coccidia infections are a significant risk factor in the health of wild quail while cryptosporidia and microsporidia may be much less significant than coccidiosis. In addition, the presence of E. hellem and E. cuniculi (known to cause opportunistic infections in humans) suggests that wild quail could serve as a reservoir for human microsporidian pathogens, and individuals with compromised or weakened immunity should probably take precautions while directly handling wild quail.

Oral Cavity and Oropharyngeal Cancer: Changing Trends in Incidence in the United States and Oklahoma.

Oral cavity cancer (OC) has steadily decreased in the United States (US) since 1973 whereas oropharyngeal cancer (OP) has increased. We analyzed OC and OP cases from the Oklahoma Central Cancer Registry and Surveillance, Epidemiology, and End Results program comparing those diagnosed from 1997-1999 to those diagnosed from 2010-2012. We compared the incidence of OC and OP cases between Oklahoma and the US and by demographic factors. We observed an increase in OP cases, but no change in OC cases in both the US and in Oklahoma, and observed some differences between Oklahoma and the US by race, gender, and age group. A possible explanation for the increasing incidence of OP cancers may be the increasing prevalence of HPV. This study highlighted the differences in temporal trends of OC and OP cancers and the importance of changing risk factors for these cancers.

Lifestyle behaviors among US cancer survivors.

PURPOSE: We describe and compare lifestyle behaviors, including smoking, physical activity, alcohol consumption, and nutrition, among cancer survivors to individuals with no cancer. METHODS: Data from the 2013 Behavior Risk Factor Surveillance System were used for this cross-sectional study. Weighted analysis was performed, and associations were examined by adjusted prevalence ratios (APRs) and 95 % confidence intervals (CIs). RESULTS: Comparing survivors to individuals with no cancer history, differences were found for a smoking quit attempt (APR 1.08; CI 1.04, 1.12), physical inactivity (APR 1.11; CI 1.07, 1.15), and binge drinking (APR 0.89; CI 0.83, 0.95). An interaction with gender was observed when examining smoking and heavy drinking. Smoking was lower (APR 0.85; CI 0.79, 0.92) among male survivors than males with no cancer history, while higher (APR 1.25; CI 1.18, 1.32) among female survivors compared to females with no cancer history. Heavy drinking (APR 0.85; CI 0.73, 0.98) was lower among male survivors than males with no cancer history, while cancer survivorship was not associated with heavy drinking among females. No differences existed for fruit and vegetable consumption or body mass index. CONCLUSIONS: US cancer survivors are not more likely than the general population to engage in all healthy lifestyle behaviors. Interventions, including improved physician communication, to reduce physical inactivity among all cancer survivors and cigarette smoking among female survivors are needed. IMPLICATIONS FOR CANCER SURVIVORS: Cancer survivors are at increased risk for comorbid conditions, and acceptance of healthy behaviors may reduce dysfunction and improve long-term health. Ultimately, opportunities exist for clinicians to promote lifestyle changes that may improve the length and quality of life of their patients.

A monoclonal antibody against the receptor for advanced glycation end products attenuates inflammatory and neuropathic pain in the mouse.

BACKGROUND: The receptor for advanced glycation end products (RAGE) is a multi-ligand receptor in the immunoglobulin superfamily. RAGE is localized throughout ascending sensory pathways (skin, peripheral nerve, dorsal root ganglion, spinal cord), and in cell types interacting with sensory neurons (endothelial cells, smooth muscle cells, monocytes and macrophages). Neuronal RAGE expression increases in pathological pain states in humans and rodents, and soluble RAGE attenuates thermal hypoalgesia in diabetic mice. The objective of the present study was to investigate whether pharmacological modulation of RAGE could attenuate mechanical allodynia in rodent pain models. METHODS: We developed an anti-RAGE monoclonal antibody (11E6) that binds to the C2 immunoglobulin domain of human RAGE, binds to mouse RAGE, and presumably to the same domain in mouse RAGE. The antinociceptive activity of 11E6 was investigated in mouse models of inflammatory (complete Freund's adjuvant) and neuropathic (chronic constriction injury of the sciatic nerve) pain. Mice were dosed intraperitoneally with 11E6 or IgG (negative control). RESULTS: Increased mechanical thresholds were observed following a single dose of 11E6 in both inflammatory and neuropathic pain models. Similar treatment with IgG did not alter nociceptive sensitivity. Repeated dosing with 11E6 significantly attenuated established mechanical hypersensitivity in a neuropathic pain model in a dose-related fashion. CONCLUSIONS: These data demonstrate that specific modulation of RAGE effectively attenuates nociceptive sensitivity associated with chronic inflammatory and neuropathic pain states.

Linking γ-aminobutyric acid A receptor to epidermal growth factor receptor pathways activation in human prostate cancer.

Neuroendocrine (NE) differentiation has been attributed to the progression of castration-resistant prostate cancer (CRPC). Growth factor pathways including the epidermal growth factor receptor (EGFR) signaling have been implicated in the development of NE features and progression to a castration-resistant phenotype. However, upstream molecules that regulate the growth factor pathway remain largely unknown. Using androgen-insensitive bone metastasis PC-3 cells and androgen-sensitive lymph node metastasis LNCaP cells derived from human prostate cancer (PCa) patients, we demonstrated that γ-aminobutyric acid A receptor (GABA(A)R) ligand (GABA) and agonist (isoguvacine) stimulate cell proliferation, enhance EGF family members expression, and activate EGFR and a downstream signaling molecule, Src, in both PC-3 and LNCaP cells. Inclusion of a GABA(A)R antagonist, picrotoxin, or an EGFR tyrosine kinase inhibitor, Gefitinib (ZD1839 or Iressa), blocked isoguvacine and GABA-stimulated cell growth, trans-phospohorylation of EGFR, and tyrosyl phosphorylation of Src in both PCa cell lines. Spatial distributions of GABAAR α1 and phosphorylated Src (Tyr416) were studied in human prostate tissues by immunohistochemistry. In contrast to extremely low or absence of GABA(A)R α1-positive immunoreactivity in normal prostate epithelium, elevated GABA(A)R α1 immunoreactivity was detected in prostate carcinomatous glands. Similarly, immunoreactivity of phospho-Src (Tyr416) was specifically localized and limited to the nucleoli of all invasive prostate carcinoma cells, but negative in normal tissues. Strong GABAAR α1 immunoreactivity was spatially adjacent to the neoplastic glands where strong phospho-Src (Tyr416)-positive immunoreactivity was demonstrated, but not in adjacent to normal glands. These results suggest that the GABA signaling is linked to the EGFR pathway and may work through autocrine or paracine mechanism to promote CRPC progression.

Expression of aldo-keto reductase family 1 member C3 (AKR1C3) in neuroendocrine tumors & adenocarcinomas of pancreas, gastrointestinal tract, and lung.

Human aldo-keto reductase family 1 member C3 (AKR1C3) was initially identified as an enzyme in reducing 5α-dihydrotestosterone (5α-DHT) to 5α-androstane-3α, 17ß-diol (3α-diol) and oxidizing 3α-diol to androsterone. It was subsequently demonstrated to possess ketosteroid reductase activity in metabolizing other steroids including estrogen and progesterone, 11-ketoprostaglandin reductase activity in metabolizing prostaglandins, and dihydrodiol dehydrogenase x (DDx) activity in metabolizing xenobiotics. AKR1C3 was demonstrated in sex hormone-dependent tissues including testis, breast, endometrium, and prostate; in sex hormone-independent tissues including kidney and urothelium. Our previous study described the expression of AKR1C3 in squamous cell carcinoma and adenocarcinoma but not in small cell carcinoma. In this report, we studied the expression of AKR1C3 in normal tissue, adenocarcinomas (43 cases) and neuroendocrine (NE) tumors (40 cases) arising from the aerodigestive tract and pancreas. We demonstrated wide expression of AKR1C3 in superficially located mucosal cells, but not in NE cells. AKR1C3-positive immunoreactivity was detected in 38 cases (88.4%) of adenocarcinoma, but only in 7 cases (17.5%) of NE tumors in all cases. All NE tumors arising from the pancreas and appendix and most tumors from the colon and lung were negative. The highest ratio of positive AKR1C3 in NE tumors was found in tumors arising from the small intestine (50%). These results raise the question of AKR1C3's role in the biology of normal mucosal epithelia and tumors. In addition, AKR1C3 may be a useful adjunct marker for the exclusion of the NE phenotype in diagnostic pathology.

Aldo-keto reductase family 1 member C3 (AKR1C3) is expressed in adenocarcinoma and squamous cell carcinoma but not small cell carcinoma.

Human aldo-keto reductase family 1 member C3 (AKR1C3) was initially identified as a critical enzyme in reducing 5α-dihydrotestosterone (5α-DHT) to 5α-androstane-3α,17ß-diol (3α-diol) and oxidizing 3α-diol to androsterone. Based on these enzymatic activities, AKR1C3 was originally named type 2 3α-hydroxysteroid dehydrogenase (HSD)/type 5 17ß-HSD. Additionally, AKR1C3 was demonstrated to be capable of metabolizing other steroids including estrogen and progesterone. Subsequently, AKR1C3 was shown to possess 11-ketoprostaglandin reductase activity in metabolizing prostaglandins and dihydrodiol dehydrogenase x (DDx) activity in metabolizing xenobiotics. Tissue distribution of AKR1C3 has been detected in both sex hormone-dependent organs such as the testis, breast, endometrium, and prostate as well as sex hormone-independent organs including the kidney and urothelium. Although prominent expression of AKR1C isozymes has been reported in human non-small cell lung carcinoma (NSCLC), the expression of AKR1C3 in small cell carcinoma of the lung has not been described. Also, the expression of AKR1C3 in normal lung has not been described. In this study, we demonstrated strong AKR1C3 immunoreactivity in bronchial epithelium but not in bronchial glands or alveolar pneumocytes. Strong AKR1C3 immunoreactivity was also demonstrated in columnar epithelium but only weak immunoreactivity in squamous epithelium of the gastrointestinal junction. Although AKR1C3 immunoreactivity was absent in small cell carcinoma of the lung, positive AKR1C3 immunoreactivity was extensively present in both adenocarcinoma and squamous cell carcinoma arising from the lung and the gastroesophageal junction. AKR1C3 may serve as an adjunct marker for differentiating small cell carcinoma from NSCLC. However, roles of AKR1C3 in adenocarcinoma, squamous cell carcinoma, and small cell carcinoma pathogenesis require further studies.

The association between narcolepsy and REM behavior disorder (RBD).

BACKGROUND AND PURPOSE: Rapid eye movement (REM) sleep Behavior Disorder (RBD) is a movement disorder associated with loss of REM-related muscle atonia and is characterized by complex, vigorous and frequently violent dream-enacting behavior during REM sleep. RBD is usually idiopathic or secondary to neurological problems such as Parkinson's disease. This study looked at the association of RBD with another sleep disorder, narcolepsy. PATIENTS AND METHODS: Seventy-eight questionnaires were sent to known narcoleptics chosen at random from those with contact details available at the center. The questionnaire addressed current narcolepsy symptoms, medication use and symptoms of RBD. Positive questionnaire results were followed up with a telephone interview. Limited polysomnography (PSG) data was also analyzed. RESULTS: Fifty-five patients responded (response rate 71%). Of these, 20 (36%) had symptoms suggestive of RBD. The typical RBD patient is an older male (mean age of onset 60.9 years, 87% male); however, in this study, females were as likely to have RBD as males, and the mean age was 41 years. Sixty-eight percent of patients who regularly experienced cataplexy and the associated symptoms of narcolepsy (sleep paralysis, hypnogogic hallucinations and automatic behavior) had RBD, compared to 14% of those who never or rarely experienced these symptoms. CONCLUSION: This study implies a stronger relationship between these disorders than a previously published figure of 7-12% This is clinically significant as RBD is a potentially distressing but readily treatable disorder. It follows that narcoleptics, especially those with cataplexy and other associated symptoms, should be questioned about symptoms of RBD and treated accordingly. Similarly, anyone presenting with RBD should be assessed for symptoms of narcolepsy, particularly if female or of a younger age group than would otherwise be expected.

Hypocretin (orexin) deficiency in narcolepsy and primary hypersomnia.

The discovery that hypocretins are involved in narcolepsy, a disorder associated with excessive daytime sleepiness, cataplexy, and unusually rapid transitions to rapid eye movement sleep, opens a new field of investigation in the area of disorders of sleep and activation. Hypocretin-1 (hcrt-1) and hypocretin-2 (hcrt-2) (also called orexin-A and orexin-B) are newly discovered neuropeptides processed from a common precursor. Hypocretin containing cells are located exclusively in the lateral hypothalamus, with widespread projections within the central nervous system. The role of the hypocretin system in other disorders causing excessive daytime sleepiness is more uncertain. This study reports the findings of a prospective study measuring cerebrospinal fluid concentrations of hypocretin-1 and hypocretin-2 in HLA DQB1*0602 positive narcolepsy with cataplexy, monosymptomatic narcolepsy, and primary hypersomnia. The results confirmed the previous observations, that hcrt-1 is deficient in narcolepsy and for the first time report very low levels of hcrt-1 in primary hypersomnia. It is also reported for the first time that there is a generalised defect in hcrt-2 transmission in all three of these clinical entities compared with controls.

Conjugated linoleic acid is synthesized endogenously in lactating dairy cows by Delta(9)-desaturase.

Conjugated linoleic acid (CLA) is a naturally occurring anticarcinogen found in milk fat and body fat of ruminants. Although CLA is an intermediate in ruminal biohydrogenation of linoleic acid, we hypothesized that its primary source was from endogenous synthesis. This would involve Delta(9)-desaturase and synthesis from trans-11 18:1, another intermediate in ruminal biohydrogenation. Our first experiment supplied lactating cows (n = 3) with trans-11 18:1 by abomasal infusion and examined the potential for endogenous synthesis by measuring changes in milk fat CLA. By d 3, infusion of trans-11 18:1 resulted in a 31% increase in concentration of cis-9, trans-11 CLA in milk fat, demonstrating that an active pathway for endogenous synthesis of CLA exists. Our second experiment examined the quantitative importance of endogenous synthesis of CLA in lactating cows (n = 3) by abomasally infusing a putative stimulator (retinol palmitate) or an inhibitor (sterculic oil) of Delta(9)-desaturase. Infusion of retinol palmitate had no influence on milk fatty acid desaturation, and yield of CLA in milk fat was not altered. However, sterculic oil infusion decreased the concentration of CLA in milk fat by 45%. Consistent with Delta(9)-desaturase inhibition, the sterculic oil treatment also altered the milk fat concentration of other Delta(9)-desaturase products as indicated by the two- to threefold increase in the ratios of 14:0 to 14:1(,) 16:0 to 16:1 and 18:0 to cis-18:1. Using changes in the ratio of 14:0 to 14:1 as an indication of the extent of Delta(9)-desaturase inhibition with the sterculic oil treatment, an estimated 64% of the CLA in milk fat was of endogenous origin. Overall, results demonstrate that endogenous synthesis of CLA from trans-11 18:1 represented the primary source of CLA in milk fat of lactating cows.

A new pathway for mitogen-dependent cdk2 regulation uncovered in p27(Kip1)-deficient cells.

BACKGROUND: The ability of cyclin-dependent kinases (CDKs) to promote cell proliferation is opposed by cyclin-dependent kinase inhibitors (CKIs), proteins that bind tightly to cyclin-CDK complexes and block the phosphorylation of exogenous substrates. Mice with targeted CKI gene deletions have only subtle proliferative abnormalities, however, and cells prepared from these mice seem remarkably normal when grown in vitro. One explanation may be the operation of compensatory pathways that control CDK activity and cell proliferation when normal pathways are inactivated. We have used mice lacking the CKIs p21(Cip1) and p27(Kip1) to investigate this issue, specifically with respect to CDK regulation by mitogens. RESULTS: We show that p27 is the major inhibitor of Cdk2 activity in mitogen-starved wild-type murine embryonic fibroblasts (MEFs). Nevertheless, inactivation of the cyclin E-Cdk2 complex in response to mitogen starvation occurs normally in MEFs that have a homozygous deletion of the p27 gene. Moreover, CDK regulation by mitogens is also not affected by the absence of both p27 and p21. A titratable Cdk2 inhibitor compensates for the absence of both CKIs, and we identify this inhibitor as p130, a protein related to the retinoblastoma gene product Rb. Thus, cyclin E-Cdk2 kinase activity cannot be inhibited by mitogen starvation of MEFs that lack both p27 and p130. In addition, cell types that naturally express low amounts of p130, such as T lymphocytes, are completely dependent on p27 for regulation of the cyclin E-Cdk2 complex by mitogens. CONCLUSIONS: Inhibition of Cdk2 activity in mitogen-starved fibroblasts is usually performed by the CKI p27, and to a minor extent by p21. Remarkably p130, a protein in the Rb family that is not related to either p21 or p27, will directly substitute for the CKIs and restore normal CDK regulation by mitogens in cells lacking both p27 and p21. This compensatory pathway may be important in settings in which CKIs are not expressed at standard levels, as is the case in many human tumors.