Adhesion of lymphoid cell lines to fibronectin-coated substratum: Biochemical and physiological characterization and the identification of a 140-kDa fibronectin receptor

Little information is available on the interaction between lymphocytes and fibronectin (fn). To gain a better understanding on this issue we examined the adhesion of 12 lymphoid cell lines, each exhibiting different phenotypic characteristics, to fn-coated substratum. Of the cell lines tested, five that adhered to fn possessed B-cell characteristics, while neither the T-cell lines nor the pre-B-cell line adhered. The physiology and biochemistry of adhesion of a B-cell line, MOPC 315, were examined in detail. Our results indicated that (1) the adhesion was a specific and time-dependent process, (2) the adhesion was temperature-dependent and inhibited by metabolic inhibitors, such as KCN and 2-deoxyglucose, (3) the presence of cycloheximide and pretreatment of cells with trypsin inhibited adhesion, (4) a 140-kDa surface protein was immunoprecipitated by anti-fn receptor antibodies, (5) the presence of divalent cations was essential for adhesion, (6) the presence of colchicine had no effect on adhesion, while cytochalasin B partially inhibited adhesion, and (7) the treatment of cells by both phorbol 12-myristate 13-acetate and calcium ionophore A23187 enhanced adhesion. In this study, we have established the interaction between lymphoid cell lines and fn. Such an interaction might play an important role in the behavior of lymphocytes in tissues.     

Cholesterol as a Risk Factor for Subarachnoid Hemorrhage: A Systematic Review

BackgroundThe role played by total cholesterol (TC) in risk for subarachnoid hemorrhage (SAH) is unclear because studies report both high and low TC each as a risk factor. We performed a systematic review to clarify associations between lipid profile and SAH.MethodsOur literature search comprised Pubmed, Scopus, and Cochrane Library databases with no language, publication year, or study type limitations. The Preferred Reporting Items for Systematic reviews and Meta-analyses (PRISMA) checklist guided our reporting. Data forms adapted from the Critical Appraisal Skills Program (CASP), and Cochrane Collaboration guidelines provided a platform for risk-of-bias evaluation. We used a random effects model to calculate pooled estimates and assessed heterogeneity with I²-statistics.ResultsOf the final 21 studies reviewed, 12 were prospective and 9 retrospective. All studies assessed TC, four assessed HDL, and none LDL in risk for SAH. Heterogeneity among all, retrospective, and Asian studies was high (I² = 79.5%, I² = 89.0%, and I² = 84.3%) and considerable in prospective (I² = 46.0%). We therefore focused on qualitative analysis and found that only two studies had a low risk of bias. According to these studies high TC increases risk for SAH in men, whereas the role of HDL remained unclear.ConclusionThe low-risk-of-bias studies suggest that elevated TC levels elevate risk for SAH in men. Due to the high prevalence of hypercholesterolemia, population attributable risk (PAR) of hypercholesterolemia may exceed the PARs of smoking and hypertension in men. Apart from diabetes and obesity, the risk-factor profile of SAH seems to resemble that of other cerebrovascular diseases, at least in men.     

Growth performance and feed conversion of Ruspolia differens on plant-based by-product diets

The use of food industry by-products in insect feeds has gained increasing attention recently. However, the understanding of how well the economically valuable edible insect Ruspolia differens (Serville) (Orthoptera: Tettigoniidae) can grow and develop with plant-based by-product feeds is currently lacking. It is important to determine the nutritional requirements, especially protein demand, of this species before developing artificial feeds for mass-rearing. We reared R. differens with four control diets and 12 plant-based by-product diets in which the major protein source came from food industry by-products, including potato-protein, barley mash, barley feed, turnip rape, a mix of broad bean and pea, and a mix of potato, carrot, and apple. We asked whether the performance (development time, survival, and weight), feed conversion, and fatty acid composition and content differed among diet treatments. Furthermore, the 12 experimental by-product diets were designed to reach six protein levels. We found that R. differens can be reared with various by-product diets, but development time, survival, and weight differed among diets. Barley feed, barley mash, and potato protein diets seem to be good options for rearing, and potato glycoalkaloids do not affect the performance of R. differens. Individuals fed on the various by-product diets also differed in their fatty acid composition and content. Increasing protein levels in diet up to 17% enhanced growth, development time, and survival, but no further enhancements were seen when fed diets with protein levels higher than this. The high protein levels decreased feed conversion rate. Our results can be valuable for designing feeds for insect mass-rearing technology. The use of food industry by-products in the diets for R. differens could increase the re-use of local resources and enhance circular economy.     

Content-Specificity in Verbal Recall: A Randomized Controlled Study

In this controlled experiment we examined whether there are content effects in verbal short-term memory and working memory for verbal stimuli. Thirty-seven participants completed forward and backward digit and letter recall tasks, which were constructed to control for distance effects between stimuli. A maximum-likelihood mixed-effects logistic regression revealed main effects of direction of recall (forward vs backward) and content (digits vs letters). There was an interaction between type of recall and content, in which the recall of digits was superior to the recall of letters in verbal short-term memory but not in verbal working memory. These results demonstrate that the recall of information from verbal short-term memory is content-specific, whilst the recall of information from verbal working memory is content-general.     

Quantitative assessment of methyl‐esterification and other side reactions in a standard propionylation protocol for detection of histone modifications

Histone modifications play an important role in regulating chromatin stability and gene expression, but to date, investigating them remains challenging. In order to obtain peptides suitable for MS‐based analysis, chemical derivatization of N‐terminus and lysine residues by propionic anhydride is commonly performed. Several side reactions (methyl‐esterification, amidation, solvolysis, overpropionylation, and missed propionylation) during propionylation protocols have been described, yet their relative abundances remain vague. Because methyl‐esterification could interfere with correct interpretation of the modification pattern, it is essential to take measures to avoid it. Here we present in‐depth quantitative analyses of methyl‐esterification and the other side reactions in a standard propionylation protocol containing methanol, and when replacing methanol with isopropanol or acetonitrile. We show that the use of alternative solvents can eliminate methyl‐esterification and that even though other side reactions are not prevented, their contribution can be kept relatively small. We also show that replacing methanol can be of importance also in other proteomics methods, such as mixed cation exchange, using methanol under acidic conditions.     

Deletion of hypoxia-inducible factor prolyl 4-hydroxylase 2 in FoxD1-lineage mesenchymal cells leads to congenital truncal alopecia

Hypoxia-inducible factors (HIFs) induce numerous genes regulating oxygen homeostasis. As oxygen sensors of the cells, the HIF prolyl 4-hydroxylases (HIF-P4Hs) regulate the stability of HIFs in an oxygen-dependent manner. During hair follicle (HF) morphogenesis and cycling, the location of dermal papilla (DP) alternates between the dermis and hypodermis and results in varying oxygen levels for the DP cells. These cells are known to express hypoxia-inducible genes, but the role of the hypoxia response pathway in HF development and homeostasis has not been studied. Using conditional gene targeting and analysis of hair morphogenesis, we show here that lack of Hif-p4h-2 in Forkhead box D1 (FoxD1)-lineage mesodermal cells interferes with the normal HF development in mice. FoxD1-lineage cells were found to be mainly mesenchymal cells located in the dermis of truncal skin, including those cells composing the DP of HFs. We found that upon Hif-p4h-2 inactivation, HF development was disturbed during the first catagen leading to formation of epithelial-lined HF cysts filled by unorganized keratins, which eventually manifested as truncal alopecia. Furthermore, the depletion of Hif-p4h-2 led to HIF stabilization and dysregulation of multiple genes involved in keratin formation, HF differentiation, and HIF, transforming growth factor β (TGF-β), and Notch signaling. We hypothesize that the failure of HF cycling is likely to be mechanistically caused by disruption of the interplay of the HIF, TGF-β, and Notch pathways. In summary, we show here for the first time that HIF-P4H-2 function in FoxD1-lineage cells is essential for the normal development and homeostasis of HFs.     

High-resolution imaging reveals a limit in spatial resolution of blood flow measurements by microspheres

Density of 15-microm microspheres after left atrial application is the standard measure of regional perfusion. In the heart, substantial differences in microsphere density are seen at spatial resolutions <5 ml, implying perfusion heterogeneity. Microsphere deposition imaging permits a superior evaluation of the distribution pattern. Therefore, fluorescent microspheres (FMS) were applied, FMS deposition in the canine heart was imaged by epifluorescence microscopy in vitro, and the patterns were observed compared with MR images of iron oxide microspheres (IMS) obtained in vivo and in vitro. FMS deposition in myocardial slices revealed the following: 1) a nonrandom distribution, with sequentially applied FMS of different color stacked within the same vessel, 2) general FMS clustering, and 3) rather large areas devoid of FMS (n = 3). This pattern was also seen in reconstructed three-dimensional images (<1 nl resolution) of FMS distribution (n = 4). Surprisingly, the deposition pattern of sequentially applied FMS remained virtually identical over 3 days. Augmenting flow by intracoronary adenosine (>2 microM) enhanced local microsphere density, but did not alter the deposition pattern (n = 3). The nonrandom, temporally stable pattern was quantitatively confirmed by a three-dimensional intermicrosphere distance analysis of sequentially applied FMS. T2-weighted short-axis MR images (2-microl resolution) of IMS revealed similar patterns in vivo and in vitro (n = 6), as seen with FMS. The observed temporally stable microsphere patterns are not consistent with the notion that microsphere deposition is solely governed by blood flow. We propose that at high spatial resolution (<2 microl) structural aspects of the vascular network dominate microsphere distribution, resulting in the organized patterns observed.     

Short-term intermittent exposure to diazoxide improves functional performance of beta-cells in a high-glucose environment

Prolonged periods of "beta-cell rest" exert beneficial effects on insulin secretion from pancreatic islets subjected to a high-glucose environment. Here, we tested for effects of short-term intermittent rest achieved by diazoxide. Rat islets were cultured for 48 h with 27 mmol/l glucose alone, with diazoxide present for 2 h every 12 h or with continuous 48-h presence of diazoxide. Both protocols with diazoxide enhanced the postculture insulin response to 27 mmol/l glucose, to 200 mumol/l tolbutamide, and to 20 mmol/l KCl. Intermittent diazoxide did not affect islet insulin content and enhanced only K(ATP)-dependent secretion, whereas continuous diazoxide increased islet insulin contents and enhanced both K(ATP)-dependent and -independent secretory effects of glucose. Intermittent and continuous diazoxide alike increased postculture ATP-to-ADP ratios, failed to affect [(14)C]glucose oxidation, but decreased oxidation of [(14)C]oleate. Neither of the two protocols affected gene expression of the ion channel-associated proteins Kir6.2, sulfonylurea receptor 1, voltage-dependent calcium channel-alpha1, or Kv2.1. Continuous, but not intermittent, diazoxide decreased significantly mRNA for uncoupling protein-2. A 2-h exposure to 20 mmol/l KCl or 10 mumol/l cycloheximide abrogated the postculture effects of intermittent, but not of continuous, diazoxide. Intermittent diazoxide decreased islet levels of the SNARE protein SNAP-25, and KCl antagonized this effect. Thus short-term intermittent diazoxide treatment has beneficial functional effects that encompass some but not all characteristics of continuous diazoxide treatment. The results support the soundness of intermittent beta-cell rest as a treatment strategy in type 2 diabetes.     

Mitochondrial Dynamics Protein Drp1 Is Overexpressed in Oncocytic Thyroid Tumors and Regulates Cancer Cell Migration

Oncocytic cell tumors are characterized by the accumulation of morphologically abnormal mitochondria in their cells, suggesting a role for abnormal mitochondrial biogenesis in oncocytic cell transformation. Little is known about the reason for the dysmorphology of accumulated mitochondria. The proteins regulating the morphology of mitochondria, the "mitochondria-shaping" proteins, can modulate their size and number; however, nothing is known hitherto about a possible involvement of mitochondrial dynamics in oncocytic cell transformation in tumors. Our aim was to assess the status of the mitochondria morphology and its role in oncocytic cell transformation. We therefore evaluated the expression pattern of the main mitochondrial fusion and fission proteins in a series of thyroid cell tumor samples, as well as in thyroid tumor cell lines, with and without oncocytic cell features. The expression of mitochondrial fusion (Opa1, Mfn1 and Mfn2) and fission (Drp1 and Fis1) proteins were evaluated by immunohistochemistry (IHC) in a series of 88 human thyroid tumors. In vitro studies, for comparative purposes and to deepen the study, were performed using TPC1 - a papillary thyroid carcinoma derived cell line—and XTC.UC1, an oncocytic follicular thyroid carcinoma-derived cell line. Both IHC and in vitro protein analyses showed an overall increase in the levels of "mitochondrial-shaping" proteins in oncocytic thyroid tumors. Furthermore, overexpression of the pro-fission protein Drp1 was found to be associated with malignant oncocytic thyroid tumors. Interestingly, genetic and pharmacological blockage of Drp1 activity was able to influence thyroid cancer cells’ migration/invasion ability, a feature of tumor malignancy. In this study we show that unbalanced mitochondrial dynamics characterize the malignant features of thyroid oncocytic cell tumors, and participate in the acquisition of the migrating phenotype.