Distribution of laminin and types IV and III collagen in fetal,infant and adult human spleens

Summary The immunohistochemical distribution of the basement membrane (BM) proteins, laminin and type IV collagen, and interstitial type III collagen was investigated in 12 fetal spleens at the 15th–38th gestational weeks (g.w.) and in spleens of 8 infants from term to 4 years. The results were compared with the distribution of the same proteins in adult human spleen. BM proteins were found to be abundantly present in the red pulp of all spleens during the whole of development. The content of type III collagen gradually decreased with advancing age and, in adult spleen, there were only occasional positively staining fibers in Billroth's cords. This finding indicates that the composition of reticular fibers in the red pulp of spleen is different from the reticular fibers elsewhere in lymphoreticular tissue. Early signs of ring fiber formation in the walls of venous sinuses were detectable at the 15th–19th g.w., although their more complete development occurred relatively late from the 36th g.w. onwards. Ring fibers contained both laminin and type IV collagen in all the investigated spleens. They never stained for type III collagen. The developing white pulp was positive for BM proteins, but showed no staining for type III collagen at the 15th g.w. At later ages, the white pulp stained similarly for both BM proteins and type III collagen.     

Handgrip strength and hand dimensions in young handball and basketball players

In handball and basketball the longer the finger length the better the accuracy of the shot or throw. All shots and throws are finished with the wrist and fingers. It can be proposed that athletes with longer fingers and greater hand surface parameters also probably have greater grip strength. The aim of this study was to investigate the influence of general body and hand-specific anthropometric dimensions on handgrip strength in boys participating in handball and basketball training. In total, 193 boys aged 10-17 years participated in this study. They were divided into 6 groups: 10-, 11-, 12-, 13-, 14-15-, and 16-17-year-olds. The body height and body mass were measured and body mass index was calculated as general anthropometric parameters. The outlines of the hands of the boys were drawn on paper with a thin marker. Three groups of hand anthropometric parameters were measured: 5 finger spans, 5 finger lengths, and 5 perimeters of the hand. Handgrip strength was measured on the dominant hand with a Lafayette dynamometer. As a rule, general anthropometric parameters determined the maximal handgrip strength more accurately than did specific hand anthropometric parameters. From the specific hand anthropometric parameters, finger lengths and perimeters of the hand significantly correlated with the maximal handgrip strength. In summary, fingers are the smallest, lightest parts of the motor apparatus, and, therefore, they represent the parts most easily deflected by force from the ball, but at the same time, finger control is especially important for the accuracy of different shots, both in handball and basketball. Thus, it is especially necessary to measure finger length and perimeters of the hand for practical reasons.     

O(2) regulates skeletal muscle progenitor differentiation through phosphatidylinositol 3-kinase/AKT signaling

Skeletal muscle stem/progenitor cells, which give rise to terminally differentiated muscle, represent potential therapies for skeletal muscle diseases. Delineating the factors regulating these precursors will facilitate their reliable application in human muscle repair. During embryonic development and adult regeneration, skeletal muscle progenitors reside in low-O(2) environments before local blood vessels and differentiated muscle form. Prior studies established that low O(2) levels (hypoxia) maintained muscle progenitors in an undifferentiated state in vitro, although it remained unclear if progenitor differentiation was coordinated with O(2) availability in vivo. In addition, the molecular signals linking O(2) to progenitor differentiation are incompletely understood. Here we show that the muscle differentiation program is repressed by hypoxia in vitro and ischemia in vivo. Surprisingly, hypoxia can significantly impair differentiation in the absence of hypoxia-inducible factors (HIFs), the primary developmental effectors of O(2). In order to maintain the undifferentiated state, low O(2) levels block the phosphatidylinositol 3-kinase/AKT pathway in a predominantly HIF1α-independent fashion. O(2) deprivation affects AKT activity by reducing insulin-like growth factor I receptor sensitivity to growth factors. We conclude that AKT represents a key molecular link between O(2) and skeletal muscle differentiation.     

Analyses of the yeast Rad51 recombinase A265V mutant reveal different in vivo roles of Swi2-like factors

The Saccharomyces cerevisiae Swi2-like factors Rad54 and Rdh54 play multifaceted roles in homologous recombination via their DNA translocase activity. Aside from promoting Rad51-mediated DNA strand invasion of a partner chromatid, Rad54 and Rdh54 can remove Rad51 from duplex DNA for intracellular recycling. Although the in vitro properties of the two proteins are similar, differences between the phenotypes of the null allele mutants suggest that they play different roles in vivo. Through the isolation of a novel RAD51 allele encoding a protein with reduced affinity for DNA, we provide evidence that Rad54 and Rdh54 have different in vivo interactions with Rad51. The mutant Rad51 forms a complex on duplex DNA that is more susceptible to dissociation by Rdh54. This Rad51 variant distinguishes the in vivo functions of Rad54 and Rdh54, leading to the conclusion that two translocases remove Rad51 from different substrates in vivo. Additionally, we show that a third Swi2-like factor, Uls1, contributes toward Rad51 clearance from chromatin in the absence of Rad54 and Rdh54, and define a hierarchy of action of the Swi2-like translocases for chromosome damage repair.     

Human Bone Marrow Mesenchymal Stem Cells Induce Collagen Production and Tongue Cancer Invasion

Tumor microenvironment (TME) is an active player in carcinogenesis and changes in its composition modify cancer growth. Carcinoma-associated fibroblasts, bone marrow-derived multipotent mesenchymal stem cells (BMMSCs), and inflammatory cells can all affect the composition of TME leading to changes in proliferation, invasion and metastasis formation of carcinoma cells. In this study, we confirmed an interaction between BMMSCs and oral tongue squamous cell carcinoma (OTSCC) cells by analyzing the invasion progression and gene expression pattern. In a 3-dimensional myoma organotypic invasion model the presence of BMMSCs inhibited the proliferation but increased the invasion of OTSCC cells. Furthermore, the signals originating from OTSCC cells up-regulated the expression of inflammatory chemokines by BMMSCs, whereas BMMSC products induced the expression of known invasion linked molecules by carcinoma cells. Particularly, after the cell-cell interactions, the chemokine CCL5 was abundantly secreted from BMMSCs and a function blocking antibody against CCL5 inhibited BMMSC enhanced cancer invasion area. However, CCL5 blocking antibody did not inhibit the depth of invasion. Additionally, after exposure to BMMSCs, the expression of type I collagen mRNA in OTSCC cells was markedly up-regulated. Interestingly, also high expression of type I collagen N-terminal propeptide (PINP) in vivo correlated with the cancer-specific mortality of OTSCC patients, whereas there was no association between cancer tissue CCL5 levels and the clinical parameters. In conclusion, our results suggest that the interaction between BMMSC and carcinoma cells induce cytokine and matrix molecule expression, of which high level of type I collagen production correlates with the prognosis of OTSCC patients.     

Size-Dependent Toxicity of Silver Nanoparticles to Bacteria,Yeast, Algae,Crustaceans and Mammalian Cells In Vitro

The concept of nanotechnologies is based on size-dependent properties of particles in the 1–100 nm range. However, the relation between the particle size and biological effects is still unclear. The aim of the current paper was to generate and analyse a homogenous set of experimental toxicity data on Ag nanoparticles (Ag NPs) of similar coating (citrate) but of 5 different primary sizes (10, 20, 40, 60 and 80 nm) to different types of organisms/cells commonly used in toxicity assays: bacterial, yeast and algal cells, crustaceans and mammalian cells in vitro. When possible, the assays were conducted in ultrapure water to minimise the effect of medium components on silver speciation. The toxic effects of NPs to different organisms varied about two orders of magnitude, being the lowest (∼0.1 mg Ag/L) for crustaceans and algae and the highest (∼26 mg Ag/L) for mammalian cells. To quantify the role of Ag ions in the toxicity of Ag NPs, we normalized the EC₅₀ values to Ag ions that dissolved from the NPs. The analysis showed that the toxicity of 20–80 nm Ag NPs could fully be explained by released Ag ions whereas 10 nm Ag NPs proved more toxic than predicted. Using E. coli Ag-biosensor, we demonstrated that 10 nm Ag NPs were more bioavailable to E. coli than silver salt (AgNO₃). Thus, one may infer that 10 nm Ag NPs had more efficient cell-particle contact resulting in higher intracellular bioavailability of silver than in case of bigger NPs. Although the latter conclusion is initially based on one test organism, it may lead to an explanation for “size-dependent“ biological effects of silver NPs. This study, for the first time, investigated the size-dependent toxic effects of a well-characterized library of Ag NPs to several microbial species, protozoans, algae, crustaceans and mammalian cells in vitro.     

Rare copy number variants observed in hereditary breast cancer cases disrupt genes in estrogen signaling and TP53 tumor suppression network

Breast cancer is the most common cancer in women in developed countries, and the contribution of genetic susceptibility to breast cancer development has been well-recognized. However, a great proportion of these hereditary predisposing factors still remain unidentified. To examine the contribution of rare copy number variants (CNVs) in breast cancer predisposition, high-resolution genome-wide scans were performed on genomic DNA of 103 BRCA1, BRCA2, and PALB2 mutation negative familial breast cancer cases and 128 geographically matched healthy female controls; for replication an independent cohort of 75 similarly mutation negative young breast cancer patients was used. All observed rare variants were confirmed by independent methods. The studied breast cancer cases showed a consistent increase in the frequency of rare CNVs when compared to controls. Furthermore, the biological networks of the disrupted genes differed between the two groups. In familial cases the observed mutations disrupted genes, which were significantly overrepresented in cellular functions related to maintenance of genomic integrity, including DNA double-strand break repair (P = 0.0211). Biological network analysis in the two independent breast cancer cohorts showed that the disrupted genes were closely related to estrogen signaling and TP53 centered tumor suppressor network. These results suggest that rare CNVs represent an alternative source of genetic variation influencing hereditary risk for breast cancer.     

Cathepsin K Is Present in Invasive Oral Tongue Squamous Cell Carcinoma In Vivo and In Vitro

Objectives

Cathepsin K, a lysosomal cysteine protease, is expressed in the tumor microenvironment (TME) of skin carcinoma, but nothing is known about cathepsin K in oral tongue squamous cell carcinoma (OTSCC). Our aim was to describe the expression of cathepsin K in invasive OTSCC in vitro and in a series of clinical cancer specimens.

Materials and Methods

OTSCC invasion in vitro was studied using invasive HSC-3 tongue carcinoma cells in 3D organotypic models. In total, 121 mobile tongue OTSCCs and 10 lymph node metastases were analyzed for cathepsin K expression. The association between cathepsin K expression and clinicopathological factors was evaluated.

Results

Cysteine protease inhibitor E64 and cathepsin K silencing significantly (p<0.0001) reduced HSC-3 cell invasion in the 3D models. Cathepsin K was expressed in a majority of carcinoma and metastatic cells, but the expression pattern in carcinoma cells did not correlate with clinical parameters. Instead, the weak expression of cathepsin K in the invasive TME front correlated with increased overall recurrence (p<0.05), and in early-stage tumors this pattern predicted both cancer recurrence and cancer-specific mortality (p<0.05 and p<0.005, respectively).

Conclusions

Cathepsin K is expressed in OTSCC tissue in both carcinoma and TME cells. Although the diminished activity and expression in aggressive tongue HSC-3 cells reduced 3D invasion in vitro, the amount of cathepsin K in carcinoma cells was not associated with the outcome of cancer patients. Instead, cathepsin K in the invasive TME front seems to have a protective role in the complex progression of tongue cancer.     

Levan Enhances Associated Growth of Bacteroides,Escherichia, Streptococcus and Faecalibacterium in Fecal Microbiota

The role of dietary fiber in supporting healthy gut microbiota and overall well-being of the host has been revealed in several studies. Here, we show the effect of a bacterial polyfructan levan on the growth dynamics and metabolism of fecal microbiota in vitro by using isothermal microcalorimetry. Eleven fecal samples from healthy donors were incubated in phosphate-buffered defined medium with or without levan supplementation and varying presence of amino acids. The generation of heat, changes in pH and microbiota composition, concentrations of produced and consumed metabolites during the growth were determined. The composition of fecal microbiota and profile of metabolites changed in response to substrate (levan and amino acids) availability. The main products of levan metabolism were acetic, lactic, butyric, propionic and succinic acids and carbon dioxide. Associated growth of levan-degrading (e.g. Bacteroides) and butyric acid-producing (e.g. Faecalibacterium) taxa was observed in levan-supplemented media. The study shows that the capacity of levan and possibly also other dietary fibers/prebiotics to modulate the composition and function of colon microbiota can be predicted by using isothermal microcalorimetry of fecal samples linked to metabolite and consortia analyses.