Culture of 3D bioprinted bone constructs requires an increased fluid dynamic stimulation.

In vitro flow-induced mechanical stimulation of developing bone tissue constructs has been shown to favor mineral deposition in scaffolds seeded with cells directly exposed to the fluid flow. However, the effect of fluid dynamic parameters, such as shear stress (SS), within 3D bioprinted constructs is still unclear. Thus, this study aimed at correlating the SS levels and the mineral deposition in 3D bioprinted constructs, evaluating the possible dampening effect of the hydrogel. Human mesenchymal stem cells (hMSCs) were embedded in 3D bioprinted porous structures made of alginate and gelatin. 3D bioprinted constructs were cultured in an osteogenic medium assessing the influence of different flow rates (0, 0.7 and 7 ml/min) on calcium and collagen deposition through histology, and bone volume (BV) through micro-computed tomography. Uniform distribution of calcium and collagen was observed in all groups. Nevertheless, BV significantly increased in perfused groups as compared to static control, ranging from 0.35±0.28 mm3, 11.90±8.74 mm3 and 25.81±5.02 mm3 at week 3 to 2.28±0.78 mm3, 22.55±2.45 mm3 and 46.05±5.95 mm3 at week 6 in static, 0.7 and 7 ml/min groups, respectively. SS values on construct fibers in the range 10-100 mPa in 7 ml/min samples were twice as high as those in 0.7 ml/min samples showing the same trend of BV. The obtained results suggest that it is necessary to enhance the flow-induced mechanical stimulation of cell-embedding hydrogels to increase the amount of mineral deposited by hMSCs, compared to what is generally reported for the development of in vitro bone constructs. STATEMENT OF SIGNIFICANCE: In this study, we evaluated for the first time how the hydrogel structure dampens the effect of flow-induced mechanical stimulation during the culture of 3D bioprinted bone tissue constructs. By combining computational and experimental techniques we demonstrated that those shear stress thresholds generally considered for culturing cells seeded on scaffold surface, are no longer applicable when cells are embedded in 3D bioprinted constructs. Significantly, more bone volume was formed in constructs exposed to shear stress values generally considered as detrimental than in constructs exposed shear stress values generally considered as beneficial after 3 weeks and 6 weeks of dynamic culture using a perfusion bioreactor.

Effects of Cyclosporine, Tacrolimus, and Rapamycin on Osteoblasts.

PURPOSE: One factor that can contribute to severe bone loss after transplantation is the direct action of immunosuppressants on bone cells. The aim of this work was to study the effects of cyclosporine (CsA), tacrolimus (FK-506), and rapamycin (RAPA) on the release of three local factors directly implicated in bone-remodeling regulation and apoptosis of human osteoblasts: interleukin (IL)-6, osteoprotegerin, and receptor activator of nuclear factor κß (RANKL). BASIC PROCEDURES: Human osteoblasts were obtained from five different patients who underwent orthopedic surgery. These cells were treated with what are considered to be a clinically high dose and an acceptable dose of each immunosuppressant-RAPA 50 ng/mL and 12 ng/mL, FK-506 20 ng/mL and 5 ng/mL, CsA 1000 ng/mL and 250 ng/mL-or vehicle. Apoptotic cell death was quantified using flow cytometry of DNA content in permeabilized, propidium iodide-stained cells. IL-6 was measured using enzyme-linked immunosorbent assay (ELISA; Quantikine Human IL6, R&D Systems, Minneapolis, Minn, United States). Messenger RNA (mRNA) expression of osteoprotegerin, RANKL, and IL-6 was measured using quantitative RT-PCR. MAIN FINDINGS: A significant increase in IL-6 (mRNA and released protein) was observed in the presence of FK-506 and RAPA. Addition of RAPA to the cultures of osteoblasts produced a significant increase in the OPG/RANKL ratio. A significant increase in osteoblast apoptosis was observed in the cells treated with FK-506 and RAPA 24 hours after the addition of immunosuppressants. CsA did not produce any significant changes in osteoblasts. PRINCIPAL CONCLUSIONS: These results suggest that an increase in osteoblast apoptosis by osteoblasts may be one of the mechanisms by which bone loss occurs after RAPA and FK-506 treatments.

Expression of bone markers and micro-CT analysis of alveolar bone during orthodontic relapse.

OBJECTIVES: To investigate biological changes in alveolar bone occurring during orthodontic relapse. MATERIALS AND METHODS: Rat maxillary first molars were moved mesially for 10 days. After orthodontic tooth movement (OTM), appliances were removed, and the molars were allowed to relapse for one, three, five, seven, 14 or 21 days. Changes in 3D morphometric parameters of bone located mesial to the first molars were evaluated by micro-CT. Total RNA was isolated from the same bone site, and real-time RT-PCR was used to measure the expression of bone formation and resorption markers. RESULTS: One day after appliance removal, the molars relapsed to a mean 73% of the achieved OTM and then steadily relapsed to 93% at 21 days. Tissue mineral density and per cent bone volume increased over the experimental period. Inversely, there was a decrease in total porosity. Gene expression of OCN, Coll-I and ALP decreased during OTM, whilst as the molars relapsed showed tended to increase. Gene expression of RANKL and TRAP increased during OTM. Changes in mRNA expression of H(+)-ATPase were minor. By 21 days post-appliance removal, the remodelling process in rats appeared to have returned to control levels. CONCLUSIONS: Bone tissue reactions on a molecular level are similar during OTM and orthodontic relapse. These findings validate the importance of immediate retention following active OTM.

Effect of TiO2 scaffolds coated with alginate hydrogel containing a proline-rich peptide on osteoblast growth and differentiation in vitro.

The aim of this study was to investigate the effect of TiO2 scaffold (SC) coated with an alginate hydrogel containing a proline-rich peptide (P2) on osteoblast proliferation and differentiation in vitro. Peptide release was evaluated and a burst release was observed during the first hours of incubation, and then progressively released overtime. No changes were observed in the cytotoxicity after 48 h of seeding MC3T3-E1 cells on the coated and uncoated TiO2 SC. The amount of cells after 7 days was higher on uncoated TiO2 SC than on alginate-coated TiO2 SC, measured by DNA content and scanning electron microscope imaging. In addition, while lower expression of integrin beta1 was detected for alginate-coated TiO2 SC at this time point, similar gene expression was observed for other integrins, fibronectin-1, and several osteoblast differentiation markers. After 21 days, gene expression of integrin beta3, fibronectin-1, osterix, and collagen-I was increased in alginate-coated compared to TiO2 SC. Moreover, increased gene expression of integrin alpha8, bone morphogenetic protein 2, interleukin-6, and collagen-I was found on P2 alginate-coated TiO2 SC compared to alginate-coated TiO2 SC. In conclusion, our results indicate that alginate-coated TiO2 SC can act as a matrix for delivery of proline-rich peptides increasing osteoblast differentiation. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2013.

Risk in dosing regimens for 25-OH vitamin D supplementation in chronic haemodialysis patients.

INTRODUCTION: 25-OH vitamin D (25-OHvitD) insufficiency or deficiency should be treated in haemodialysis (HD) patients, although the 25-OHvitD target, drug or dosing regimens are unclear. AIMS: To describe factors associated with 25-OHvitD levels in HD patients and to assess the effect of three dosing regimens to supplement 25-OHvitD (calcifediol) on serum calcium (Ca), phosphate (P), parathyroid hormone (PTH), 25-OHvitD and 1,25-OHvitD. METHODS: Two hundred and seventeen patients from three HD units were studied. Demographic and biochemical data were collected at baseline. Two different 25-OHvitD assays were used. One hundred and sixty-seven patients were treated with various calcifediol dosing regimens. The same biochemical determinations were repeated after 3 months of treatment. RESULTS: At baseline, 12.9% of patients had 25-OHvitD <10 ng/ml. In multivariate linear regression, the season (lower in winter) and the assay method were determinants of 25-OHvitD concentration. Following calcifediol supplementation, 25-OHvitD, calcium and phosphate increased, while PTH diminished with statistical significance. After treatment, there were positive correlations between 25-OHvitD and Ca (r = 0.28, p < 0.0001) or 1,25-OHvitD (r = 0.75, p < 0.0001) that were not observed in the baseline dataset. High concentrations of post-treatment 25-OHvitD were associated with higher 1,25-OHvitD levels. Calcemia increased more in those treated with concomitant active vitamin D or those having suppressed baseline PTH, while PTH decreased more in those having above-target PTH levels. CONCLUSIONS: Standardisation of methods to determine 25-OHvitD blood levels is needed. In HD patients, calcifediol increased 25-OHvitD, calcemia and phosphatemia and lowered PTH. Caution should be exercised with the higher calcifediol dosing regimens, especially in patients with suppressed PTH or on vitamin D receptor activators.

Effect of alginate hydrogel containing polyproline-rich peptides on osteoblast differentiation.

Polyproline-rich synthetic peptides have previously been shown to induce bone formation and mineralization in vitro and to decrease bone resorption in vivo. Alginate hydrogel formulations containing these synthetic peptides (P2, P5, P6) or Emdogain® (EMD) were tested for surface coating of bone implants. In an aqueous environment, the alginate hydrogels disclosed a highly compact structure suitable for cell adhesion and proliferation. Lack of cytotoxicity of the alginate-gel coating containing peptides was tested in MC3T3-E1 cell cultures. In the present study, relative mRNA expression levels of integrin alpha 8 were induced by P5 compared to untreated alginate gel, and osteopontin mRNA levels were increased after 21 days of culture by treatment with synthetic peptides or EMD compared to control. Further, in agreement with previous results when the synthetic peptides were administered in the culture media, osteocalcin mRNA was significantly upregulated after long-term treatment with the formulated synthetic peptides compared to untreated and EMD alginate gel. These results indicate that the alginate gel is a suitable carrier for the delivery of synthetic peptides, and that the formulation is promising as biodegradable and biocompatible coating for bone implants.

[Physical compatibility of sodium bicarbonate with other drugs often administered in the intensive care unit]. / Compatibilidad física del bicarbonato sódico con fármacos de uso frecuente en la unidad de cuidados intensivos.

INTRODUCTION: Acid-base disorders are associated with many diseases of the critically ill patient. Early treatment with sodium bicarbonate of these metabolic disorders is required for their normalization. This is an alkaline electrolyte solution administered by continuous or intermittent infusion. Its incompatibility due to its alkaline pH has been described, as it produces insoluble carbonate precipitation and causes carbon dioxide gas when mixed with acidic solutions. MATERIAL AND METHOD: An in vitro experimental study was performed. Bicarbonate was mixed with 13 drugs simulating Y-site administration. We combined 5 ml bicarbonate with 5 ml of every drug, at highest daily concentration used. The samples were visually examined to detect color changes, cloudiness, precipitation or gas formation, pH measurement and spectrophotometric analysis at 450 nm and 620 nm. The samples were evaluated at 0, 15, 30, 60 and 120 minutes. The compatibility criteria were absence of visual changes, pH changes<0.5 and variability of absorbance <0.01. RESULTS: We studied each drug individually and mixed with bicarbonate with 27 samples, and 135 measurements were performed. The incompatibilities did not always produce visual changes. Knowing the pH of drugs does not guarantee the compatibility of the mixture. Nitroglycerin with pH 4 is compatible with bicarbonate. Thiopental with pH 11 makes precipitation. Higher absorbances showed color changes, cloudiness and precipitation. CONCLUSIONS: Bicarbonate is physically compatible with esmolol, furosemide, heparin, insulin, morphine, nimodipine, nitroglycerin and urapidil and incompatible with amiodarone, cisatracurium, haloperidol, midazolam and thiopental.

Daily or monthly ibandronate prevents or restores deteriorations of bone mass, architecture, biomechanical properties and markers of bone turnover in androgen-deficient aged rats.

AIM: The aim of this study was to investigate the effects of the bisphosphonate ibandronate (IBN) in a male osteoporosis animal model. METHODS: Two studies were performed in 9-month-old orchidectomised (ORX) or sham-operated rats. In prevention study, subcutaneous IBN was administered daily (1 µg/kg) or monthly (28 µg/kg every 28 days) starting on day of surgery for 5 months. In treatment study, the same treatment started 6 months after ORX. After sacrifice, bone analyses by dual-energy X-ray absorptiometry, 3-dimensional micro-computed tomography, and 3-point bending were performed in femora or vertebrae. Serum tartrate-resistant acid phosphatase 5b (TRAP-5b) and aminoterminal propeptide of collagen I (PINP) were analysed for resorption and osteocalcin (BGP) for bone formation. RESULTS: In both studies, ORX resulted in significant femoral and vertebral bone loss and microarchitectural deterioration after 5 months of ORX, and became more pronounced after 11 months. Biomechanical strength was also decreased. Serum levels for TRAP-5b and BGP increased while PINP levels were reduced or unchanged. Both daily and monthly IBN prevented or even restored ORX-induced changes in both studies, with the intermittent regimen showing a improvement in efficacy with respect to many of the biomechanical parameters.

[Physical compatibility of furosemide in intravenous mixtures for continuous perfusion]. / Compatibilidad visual y física de la furosemida en mezclas intravenosas para perfusión continua.

INTRODUCTION: Patients in the intensive care units simultaneously receive concomitantly many drugs, with limited venous accesses. Thus, several different perfusions must be administered jointly through the same catheter. Furosemide is a solution with no buffer capacity that is easily precipitated. In spite of the recommendation on its data sheet stating that it should not be used with other drugs, studies found in the bibliography suggest that it has physicochemical compatibility with other drugs. However, the concentrations studied do not always coincide with those used in the clinical practice. OBJECTIVES: To study and verify the physical compatibility of furosemide in intravenous mixture. MATERIAL AND METHOD: An experimental study in vitro, in which furosemide was mixed with 12 drugs at a 1:1 proportion was performed. A total of 40 samples were obtained and the following variables were evaluated at different intervals, 0, 15, 30, 60 and 120 min: ph of mixture, color changes, presence of turbidity and precipitation. To do so, visual observation procedures, ph measurements and spectrophotometer absorption at 450 and 620 nm were used. RESULTS: Forty samples, 13 simples, 12 doubles and 15 triples, were obtained. Those mixtures that did not show physical changes, pH variation and absorption variation were considered compatible. CONCLUSION: Furosemide is physically compatible with bicarbonate solution, heparin, insulin, morphine and nitroglycerin and incompatible with amiodarone, cisatracurium, haloperidol, midazolam and urapidil.

Differentiation of Y chromosome in the X(1)X(1)X(2)X(2)/X(1)X(2)Y sex chromosome system of Hoplias malabaricus (Characiformes, Erythrinidae).

Six populations of Hoplias malabaricus from the Lower Paranapanema River were studied and showed 2n = 40 with metacentric/submetacentric chromosomes for females and 2n = 39 with m/sm chromosomes for males, thereby revealing a multiple sex chromosome system of the X(1)X(1)X(2)X(2)/X(1)X(2)Y type. The Y chromosome was the largest metacentric chromosome in all karyotypes. The heterochromatic blocks were located in a pericentromeric region of all pairs and in telomeric regions in some other pairs. A conspicuous pericentromeric C-band was detected in a single pair, equivalent to the X(1) chromosome. No markers on X(2) chromosomes were detected, but they could be identified by their size. The females displayed the same distribution of C and chromomycin A(3) bands and 18S rDNA sites among the populations. However, the males exhibited differences in both number and position of Giemsa C-bands on the Y chromosome, which were also evident after Chromomycin A(3) banding and FISH with the 18S rDNA probe. These cytogenetic tools allowed for the identification of possible mechanisms involved in the differentiation and evolution of the Y chromosome.

[Perception of privacy in patients hospitalized in an Intensive Care Unit]. / Percepción de la intimidad en pacientes hospitalizados en una Unidad de Cuidados Intensivos.

INTRODUCTION: The Intensive Care Unit (ICU) is an impersonal and technologically invasive setting. The rules and framework of the unit favor a lack of privacy, an inherent fact for every kind of hospitalization. The objectives of the study are to know the experience of patients admitted to the ICU in relation to the perception of privacy and its impact on their own experience. MATERIAL AND METHODS: Phenomenological qualitative methodology. The participants were patients admitted to the ICU of Son Dureta University Hospital in Palma de Mallorca, who were older than 18 with a minimum stay of 48 hours and who had signed an informed consent. Data collection. By exhaustive semi-structured interviews. Content analysis. RESULTS: The perception of privacy in patients admitted to the ICU is described in relationship with surrounding physical space, family setting and privacy autonomy. The patients evaluated the flexibility of rules, professionalism of the nursing staff and the need for family support when the situation is more stable. CONCLUSION: To prevent the quality of nursing cares from only depending on their technological knowledge and ability, but also depends on the humane side of caring, it is extremely important to consider the different elements that mould the experience of being admitted to the ICU and respect of the privacy of each person.

Well-being, appraisal, and coping in Latina and Caucasian female dementia caregivers: findings from the REACH study.

While there has been considerable interest in studying ethnically diverse family caregivers, few studies have investigated the influence of dementia caregiving on Latino families. The current study includes participants from two sites of the REACH (Resources for Enhancing Alzheimer's Caregiver Health) project to compare well-being, appraisal, and religiosity by ethnicity, with specific attention to levels of acculturation. Latina (n = 191) and Caucasian female (n = 229) dementia family caregivers from two regions of the United States (Miami, Florida and Northern California) were compared at baseline on demographics, care recipient characteristics, mental and physical health, and psychosocial resources, including appraisal style and religiosity. Latina caregivers reported lower appraisals of stress, greater perceived benefits of caregiving, and greater use of religious coping than Caucasian caregivers. The relationship of these variables to level of acculturation for the Latina caregivers was also explored. Implications of these results for psychosocial interventions with Latino and Caucasian family caregivers are discussed.

An evaluation of the NINCDS-ADRDA neuropsychological criteria for the assessment of Alzheimer's disease: a confirmatory factor analysis of single versus multi-factor models.

Neuropsychological test batteries are frequently used to assess the nature and severity of cognitive deficits among patients with early Alzheimer's Disease (AD) and related disorders. The NINCDS-ADRDA criteria are among the most widely used guidelines to diagnose dementia (McKhann et al.,1984). These criteria specify eight distinct areas of neuropsychological function that should be evaluated in patients with suspected cognitive impairment. Recent studies have suggested that neuropsychological deficits observed in AD may be explained by a single general factor related to memory deficits or to executive dysfunction. In contrast, the results of other investigations have indicated that multiple qualitatively different factors underlie cognitive abilities in AD. In the present study, we used confirmatory factor analysis to examine the structure of cognitive abilities in AD and to assess the extent to which single and multiple ability factors accurately represent neuropsychological test data obtained from patients with AD. Results indicated that the NINCDS-ADRDA model fit the data better than a single factor model. However, a more parsimonious model specifying memory, verbal abilities, visuospatial skills, executive function, and higher as well as lower functional activities of daily living fit the data better than the NINCDS-ADRDA model. These results have important theoretical and practical implications for diagnostic evaluation.

[The age of onset of alcohol-related problems as a prognostic factor in abstinence in patients admitted to a general hospital]. / La edad de inicio de los problemas por el alcohol como factor pronóstico de abstinencia en pacientes ingresados en el Hospital General.

Seventy-three inpatients of a general hospital were studied prospectively. Detection of alcohol problems was done by non-psychiatrist physicians. The psychiatrist, after evaluating the problem recommended an outpatient centre for treatment. One year after this 15% of them remained abstinent. Age onset of alcohol-related problems correlated positively with abstinence days number. Abstinence was associated with an late age onset of alcohol-related problems and contacting with the outpatient center. Thirty-seven years age was the cutoff point with more specificity and sensibility. Hypothesis for this results are discussed.

A qualitative analysis of semantic intrusive errors in Alzheimer's disease.

Intrusive or perseverative errors as evidenced on tests of memory have been found to occur with greater frequency in patients with Alzheimer's Disease (AD) relative to other disorders and are thought to reflect the cholinergic deficits associated with AD. A particular type of intrusive error, semantic intrusions, has been found to discriminate AD from multi-infarct dementia and normal elderly controls. However, it has been suggested that such errors actually reflect random responding as the result of the disinhibition often observed in AD patients. The present study compared the frequency and types of intrusive errors from a sample of mildly, moderately, and more severely impaired AD patients who had undergone the Fuld Object Memory Evaluation with the "guesses" of elderly controls. Results of this study indicate that the semantic intrusions of AD patients differ both in type and frequency from the guesses of the control subjects.

Neuropsychological test performance and prediction of functional capacities among Spanish-speaking and English-speaking patients with dementia.

Neuropsychological measures have been widely used by clinicians to assist them in making judgments regarding a cognitively impaired patient's ability to independently perform important activities of daily living. However, important questions have been raised concerning the degree to which neuropsychological instruments can predict a broad array of specific functional capacities required in the home environment. In the present study, we examined 127 English-speaking and 56 Spanish-speaking patients with Alzheimer's disease (AD) and determined the extent to which various neuropsychological measures and demographic variables were predictive of performance on functional measures administered within the clinical setting. Among English-speaking AD patients, Block Design and Digit-Span of the WAIS-R, as well as tests of language were among the strongest predictors of functional performance. For Spanish-speakers, Block Design, The Mini-Mental State Evaluation (MMSE) and Digit Span had the optimal predictive power. When stepwise regression was conducted on the entire sample of 183 subjects, ethnicity emerged as a statistically significant predictor variable on one of the seven functional tests (writing a check). Despite the predictive power of several of the neuropsychological measures for both groups, most of the variability in objective functional performance could not be explained in our regression models. As a result, it would appear prudent to include functional measures as part of a comprehensive neuropsychological evaluation for dementia.

Deterioration of functional capacities in Alzheimer's disease after a 1-year period.

There is a paucity of data regarding Alzheimer's disease (AD) patients' longitudinal deterioration in the ability to conduct numerous activities required for daily living. In this study, 52 patients with AD were assessed at baseline and at a 1-year follow-up using the Direct Assessment of Functional Status (DAFS) scale, an objective, well-validated measure of a broad spectrum of functional capacities that is administered within the clinical setting. An important finding was that the level of initial performance on each of the 11 functional tasks measured did not relate to the degree of functional decline in that particular area. Communication skills, such as using the telephone (deterioration among 35.4% of the patients) and preparing a letter for mailing (deterioration among 32.7%), showed the most frequent deterioration among patients upon follow-up. More than half of the AD patients studied demonstrated impairment on one or both of these measures. The pattern of findings indicates that many subtests of the DAFS were sensitive to functional decline after a 1-year period and that the scale has utility in objectively establishing longitudinal patterns of deterioration.

Caregiving for Hispanic Alzheimer's disease patients: understanding the problem.

The authors analyze aspects of caregiving for Hispanic Alzheimer's disease (AD) patients. A review of the literature on caregiving and ethnicity and data concerning caregiving for AD patients in different Hispanic groups are presented. The presence of ethnicity-related differences in caregiving for Hispanic AD patients is confirmed. These differences, however, appear to be present largely in certain areas, such as household composition, distribution of caregiving roles, help-seeking behavior, and site of caregiving. The authors consider areas that need to be addressed in future research and specific modifications to current caregiving models for the study of AD patients in the Hispanic community.

Daughters caregiving for Hispanic and non-Hispanic Alzheimer patients: does ethnicity make a difference?

This study assessed Cuban-American Hispanic and White non-Hispanic daughters who were major caregivers for their mothers suffering from Alzheimer's Disease. Although patients in both ethnic groups did not differ in their level of cognitive and functional impairment, Cuban-American Hispanic patients were significantly more likely to be living in their daughters' homes while the White non-Hispanic patients resided in institutional settings. Caregivers were equivalent in their knowledge and utilization of community services, but Cuban-American daughters were significantly more aware of financial aid resources. Cuban-American patients were significantly more depressed than their White non-Hispanic counterparts with daughters showing similar but nonsignificant trends. The impact of cultural factors on caregiving is discussed.

Attention deficit disorder in children at risk for anxiety and depression.

The comorbidity between attention deficit disorder (ADD) and anxiety and/or depressive disorders was examined in the children of parents with panic disorder, major depressive disorder, or with no diagnosis. A child received a diagnosis by a self-report, parent report, and by consensus, using a best estimate procedure. The prevalence rates of ADD were significantly greater in offspring of parents with depressive and panic disorder by the parents' report and in children of depressed parents by consensus. A significant relationship between ADD and anxiety and/or depression was found for parent, child, and consensus diagnoses. Higher rates of ADD were reported by children (1% versus 13%), parents (8% versus 31%), and in the consensus diagnoses (13% versus 29%) when anxiety and/or depression was present. These results suggest that in children referred for evaluation of ADD, the possibility of a primary anxiety or depressive disorder should be considered.