Cryopreserved mouse pancreatic acinar cells from long-term explant outgrowth cultures maintain their secretory phenotype after thawing

Background/objectivesWe recently reported an explant outgrowth culture method for obtaining functionally competent mouse pancreatic acinar cells for long-term in vitro purposes. The aim of the present study was to explore the possibility of cryostoring these cells without loss of functional differentiation.MethodsAcinar cells prepared by the explant outgrowth method were cryopreserved using a DMSO-based protocol and stored in liquid nitrogen for 4 weeks. The following characteristics were compared in cryopreserved and parallel non-frozen cell preparations: cell viability and recovery, amylase content in viable cells before culture, basal and stimulated amylase release in culture and the ability of the cells to form glandular structures in Matrigel.ResultsImmediate post-thaw viability of the cells was similar to that of freshly isolated cells. Approximately 53% of viable cells frozen were recovered after thawing. Intracellular amylase content was identical in frozen and non-frozen cells. Cryopreserved cells maintained their ability to secrete amylase and to respond to caerulein stimulation in 4-day secondary cultures. They also were observed to form amylase-expressing glandular structures in three-dimensional cultures in Matrigel in a similar manner as non-frozen cells.ConclusionsThis study shows that pancreatic acinar cells can be cryopreserved for long-term storage in liquid nitrogen without dedifferentiation. Successful cryopreservation helps to refine the experimental use of primary acinar cells by enabling their banking for on-demand utilization.     

Sex differences in striatal presynaptic dopamine synthesis capacity in healthy subjects.

BACKGROUND: There are sex differences in the clinical features of several neuropsychiatric illnesses associated with dopamine dysfunction. The effects of sex on brain dopaminergic function have been sparsely studied in human subjects using modern imaging techniques. We have previously reported that the apparent affinity of [(11)C]raclopride for striatal D(2) dopamine receptors in vivo is lower in women than in men, whereas D(2) receptor density is not different. This finding indirectly suggests that women have a higher synaptic concentration of dopamine in the striatum. We explored further the basis of this phenomenon in an independent study and hypothesized that striatal presynaptic dopamine synthesis capacity would also be elevated in women. METHODS: A total of 23 healthy men and 12 healthy women (age range 20-60 years) were studied using positron emission tomography and [(18)F]fluorodopa. RESULTS: Women had significantly higher striatal [(18)F]fluorodopa uptake (Ki values) than men. The difference was more marked in the caudate (+26%) than in the putamen (+12%). In addition, there was a negative correlation between striatal [(18)F]fluorodopa Ki values and age in men but not in women. CONCLUSIONS: The results further substantiate sex differences in striatal dopaminergic function in humans. This finding may be associated with sex differences in vulnerability and clinical course of neuropsychiatric disorders with dopaminergic dysregulation, e.g., schizophrenia, alcohol dependence, and Parkinson's disease.     

Intrauterine hypoxia as a cause of hearing impairment in children

Among more than 10,000 mothers giving birth during 1974–1977, there were 20 known cases of severe placental insufficiency in connection with hypertensive disease during pregnancy. Well-documented signs of intrauterine hypoxia were observed in all the cases. In 50% of the fetuses intrauterine growth retardation was found to be below the 10th percentile curve. In most cases the baby was delivered by means of a caesarean section. The hearing level of these infants was examined at the age of 2–5 years. In these 20 children there were no signs of sensorineural hearing loss; however, because of the size of the population, the conclusions drawn from this thorough work must be tentative. It seems that with good maternity care and thorough treatment hearing loss due to intrauterine hypoxia can be prevented.     

Caring from the point of view of a Finnish mental health nurse: a life history approach

Finnish mental health care went through a major change in the 1980s. Its institutions were being run down and there was a shift to a caring system that stresses non-institutional practices. The schooling and work of the mental health nurse had to adapt to the new situation. The purpose of my study was to describe and analyse the mental health nurse's work, i.e. caring, as it is experienced by a nurse in the preparation of long-term psychiatric patients for non-institutional care. The data consist of repeated interviews with Vuokko, who has been a mental health nurse for about 30 years and is now approaching her retirement. In the light of the data, the central features of caring are caring about and taking an overall responsibility for the patient, and teaching the patient to cope with everyday life. Caring is based on practical knowledge, the essential elements of which are intimacy and feelings.     

Serum antibody response to periodontal pathogens and herpes simplex virus in relation to classic risk factors of cardiovascular disease

BACKGROUND: Increasing evidence links chronic infections, especially burden of several infections, with increased risk for cardiovascular diseases (CVD). We studied joint immune response against two major periodontal pathogens and herpes simplex virus (HSV) in relation to established risk factors of CVD. METHODS: Serum antibody levels to HSV, Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis were determined by ELISA. The study included 1107 subjects, 734 from Finland and 373 from Russia. RESULTS: Combined antibody response to periodontal pathogens was associated inversely (OR, 95% CI) with high-density lipoprotein (HDL) cholesterol concentration (beta = 0.35; 0.20, 0.60; P < 0.001) and directly with HSV antibody quartiles: compared with the first quartile, ORs (95% CI) for quartiles 2-4 were 1.43 (0.88-2.32), 1.74 (1.07-2.82), and 1.89 (1.18-3.02), respectively (P for trend <0.001), after adjusting for age, gender, area, education, smoking, BMI, alcohol, triglycerides, and number of teeth. In linear regression analysis, the 3-pathogen antibody score (comprising antibody levels against periodontal pathogens and HSV) was inversely associated with HDL cholesterol concentration (beta = -0.067/1 mmol/l; -0.235, -0.018; P < 0.05). CONCLUSIONS: HSV infection may promote infection by periodontal pathogens. Furthermore, the infectious burden comprising HSV and periodontitis may increase the risk for CVD by clearly decreasing HDL cholesterol concentrations.     

How much vitamin D3 do the elderly need?

BACKGROUND: Vitamin D insufficiency poses a problem in many parts of the world, the elderly being an especially vulnerable group. This insufficiency results from an inadequate amount of sunshine and a low dietary intake of vitamin D. Typically, insufficiency is accompanied with high intact parathyroid hormone, (S-iPTH) concentrations. AIMS OF THE STUDY: We studied how serum 25-hydroxy vitamin D (S-25-OHD) concentrations respond to different doses of vitamin D3 supplementation. Secondly to determine the smallest efficient dose to maintain serum 25-OHD concentration above the insufficiency level. We also studied which dose would be efficient in decreasing S-iPTH concentration in these subjects. SUBJECTS AND METHODS: Forty-nine 65- to 85-year-old women participated. The women were randomly assigned into one of four groups receiving 0 (placebo), 5, 10 or 20 microg of vitamin D3 daily for 12 weeks. Fasting morning blood was drawn at the beginning of the study, and thereafter every second week. Calciotropic variables were assessed from serum and urine samples. RESULTS: The S-25-OHD concentration increased significantly (p < 0.001) in all supplemented groups [5 microg: by 10.9 (8.5) nmol/L, 10 microg: by 14.4 (6.9) nmol/L, 20 microg: by 23.7 (11.9) nmol/L], whereas it decreased in the placebo group by 8.3 (13.2) nmol/L. Equilibrium in S-25-OHD concentration was reached in all groups after 6 weeks of supplementation at 57.7 (8.9) nmol/L, 59.9 (8.9) nmol/L and 70.9 (8.9) nmol/L in the groups with increasing vitamin D supplementation. The dose-response to supplementation decreased with increasing vitamin D status at baseline, r = -0.513, p = 0.002. S-iPTH tended to decrease in those with highest dose response to supplementation. CONCLUSIONS: A clear dose response was noted in S-25-OHD to different doses of vitamin D3. The recommended dietary intake of 15 microg is adequate to maintain the S-25-OHD concentration around 40-55 nmol/L during winter, but if the optimal S-25-OHD is higher than that even higher vitamin D intakes are needed. Interestingly, subjects with lower vitamin D status at baseline responded more efficiently to supplementation than those with more adequate status.     

Comparison of oxycodone pharmacokinetics after buccal and sublingual administration in children

OBJECTIVE: We evaluated and compared the pharmacokinetics of two oral administration routes of oxycodone parenteral liquid (10 mg/mL)--single buccal and sublingual administration--in 30 generally healthy awake children, aged 6-91 months. METHODS: Two groups of children undergoing inpatient surgery were enrolled. In a randomised fashion, children received a single dose of oxycodone 0.2 mg/kg buccally (n = 15) or sublingually (n = 15). Regular blood samples were collected for up to 12 hours, and plasma was analysed for oxycodone, oxymorphone and noroxycodone using gas chromatography-mass spectrometry. RESULTS: Bioavailability was similar after administration at the two instillation sites. The area under the plasma concentration-time curve from time zero extrapolated to infinity (AUCinfinity) was 2400-8000 ng x min/mL (median 4200 ng x min/mL) in the buccal group and 2700-7900 ng x min/mL (median 5500 ng x min/mL) in the sublingual group. After buccal administration, maximum plasma concentration (Cmax) was 5.4-39 ng/mL (16 ng/mL) after buccal and 5.5-42 ng/mL (22 ng/mL) after sublingual administration. Twelve of the 15 children in both groups reached the oxycodone analgesic concentration of 12 ng/mL, which was sustained for 43-209 minutes (median 160 minutes) in the children with buccal oxycodone and for 32-262 minutes (median 175 minutes) in the children with sublingual oxycodone. The terminal elimination half-lives were closely similar in the two groups: 104-251 minutes (median 140 minutes) in the buccal group and 110-190 minutes (150 minutes) in the sublingual group. CONCLUSION: The results of this study show that in young children the absorption of oxycodone is similar after buccal and sublingual instillation.     

Noninvasive imaging for evaluation of the systemic delivery of capsid-modified adenoviruses in an orthotopic model of advanced lung cancer

BACKGROUND: Variable expression of the coxsackie and adenovirus receptor (CAR) has limited gene transfer efficacy to many types of tumors. Consequently, tropism-modified adenoviruses have been developed for enhanced infectivity. To the authors' knowledge, targeting approaches for nonsmall cell lung cancer (NSCLC) have not been comprehensively evaluated. The current hypothesis was that modified adenoviruses could be used for increasing gene transfer to and killing of NSCLC cells in vitro and in vivo. METHODS: Ten NSCLC cell lines were analyzed to represent the different NSCLC histologies. Because clinical tumors may differ from established cell lines, 6 clinical specimens fresh from patients were analyzed. For in vivo studies, a novel orthotopic murine model of advanced lung cancer was developed. Because tumor response is difficult to quantitate in orthotopic models, noninvasive imaging of green fluorescent protein (GFP) was utilized as a surrogate for tumor size measurements. RESULTS: Adenoviruses whose capsids were modified with RGD-4C, the serotype 3 knob, or polylysine displayed increased gene transfer to NSCLC cell lines and clinical samples in comparison to serotype 5 viruses. Conditionally replicating oncolytic adenoviruses (CRAds) with the same modifications showed enhanced therapeutic efficiency in vitro and in vivo. The median survival of mice treated with Ad5.pK7-Delta24 or Ad5-Delta24RGD increased 37% (P<.01). GFP imaging allowed noninvasive individualized detection of response and recurrence. CONCLUSIONS: Targeting of adenovirus to heterologous receptors can improve killing of NSCLC cells. Utilization of clinical samples and an orthotopic model of advanced lung cancer may provide clinically relevant translational data.     

Amplification of the urokinase gene and the sensitivity of prostate cancer cells to urokinase inhibitors

OBJECTIVES: To evaluate the frequency of the urokinase-type plasminogen activator (uPA) gene amplification and the sensitivity of prostate cancer cells to uPA inhibition, as we previously found one hormone-refractory prostate tumour with high-level amplification of the uPA (alias PLAU) gene, and also showed that a uPA inhibitor, amiloride, can effectively reduce the invasion potential of the PC-3 prostate cancer cell line. MATERIALS AND METHODS: Sixty-three locally recurrent hormone-refractory tumours and 78 hormone-refractory metastases from 29 patients who died from prostate cancer were analysed for uPA gene-copy number using fluorescence in situ hybridization. The Matrigel invasion assay was used to study the influence of uPA inhibitors on the invasive potential of prostate cancer cell lines. RESULTS: Of the locally recurrent hormone-refractory tumours, 21% had an increased copy number of uPA, but no high-level amplifications were found; 31% of the metastases had increased copy number and one high-level amplification of the uPA. Matrigel invasion assays with two specific uPA inhibitors, B428 and p-aminobenzamidine, showed that invasion of a prostate cancer cell line containing uPA gene amplification was inhibited by these small-molecule uPA inhibitors, while invasion of prostate cell lines without uPA gene amplification were not. CONCLUSION: These results suggest that selective inhibition of the uPA pathway in individuals whose tumours contain uPA gene amplification may provide therapeutic benefit.