Mechanical coupling of supracellular stress amplification and tissue fluidization during exit from quiescence

Cellular quiescence is a state of reversible cell cycle arrest that is associated with tissue dormancy. Timely regulated entry into and exit from quiescence is important for processes such as tissue homeostasis, tissue repair, stem cell maintenance, developmental processes, and immunity. However, little is known about processes that control the mechanical adaption to cell behavior changes during the transition from quiescence to proliferation. Here, we show that quiescent human keratinocyte monolayers sustain an actinomyosin-based system that facilitates global cell sheet displacements upon serum-stimulated exit from quiescence. Mechanistically, exposure of quiescent cells to serum-borne mitogens leads to rapid amplification of preexisting contractile sites, leading to a burst in monolayer tension that subsequently drives large-scale displacements of otherwise motility-restricted monolayers. The stress level after quiescence exit correlates with the level of quiescence depth at the time of activation, and a critical stress magnitude must be reached to overcome the cell sheet displacement barrier. The study shows that static quiescent cell monolayers are mechanically poised for motility, and it identifies global stress amplification as a mechanism for overcoming motility restrictions in confined confluent cell monolayers.

Quiescence refers to a state of cell cycle arrest in which cells are retained in a standby mode, ready to re-enter the cell cycle upon activation by a given physiological stimuli. The pool of quiescent cells in the human body is typically represented by tissue-specific stem and progenitor cells, naive immune cells, fibroblasts, and epithelial cells (1, 2). In addition, certain cancer cells have the ability to evade cancer therapy by entering a dormant quiescence-like state (1, 2). Accordingly, careful regulation of entry into and exit out of quiescence is important for several physiological processes such as tissue homeostasis and repair, stem cell maintenance, immunity, reproduction, and development (1, 2).During homeostasis, the balance between quiescent and proliferating cells is controlled by constituents of the microenvironment such as soluble factors, extracellular matrix components, blood vessels, and neighboring cells. On the other hand, during episodes that require extensive tissue renewal and remodeling, for example after injury, coordinated stimulation of quiescent cells into proliferation is facilitated by increased exposure to blood-borne and cell-secreted mitogens through local inflammatory responses such as increased blood flow, increased vascular permeability (vasodilation), and immune cell recruitment (3, 4). Accordingly, a commonly used methodology for studies of quiescence in cultured mammalian cells involves consecutive treatments with serum-free and serum-containing growth medium (1).Quiescent cells are required to maintain a high level of preparedness in order to facilitate rapid activation of specialized cell functions once cell division is stimulated. In agreement with this, quiescent stem cells and naive immune cells have been shown to possess multiple epigenetic and posttranslation mechanisms that facilitate the rapid expression of linage-specific genes following stimulation of quiescence exit (2, 5–14). However, little is known about mechanical forces that facilitate adaptation to cell cycle–activated behaviors.Quiescence exit is frequently associated with activation of cell motility. For example, quiescent stem and naive immune cells migrate out of their niches in response to cell cycle activation in order to support tissue homeostasis, repopulate injured tissue, or to perform immune surveillance at distal locations (15–18). In addition, reawakening of dormant quiescent cancer cells can cause tumor relapse and formation of metastases years after remission (19). In multilayered epithelial tissue, like the skin, exit from quiescence during homeostasis is associated with lateral migration to suprabasal regions, while skin injury evokes massive reawakening of basally localized keratinocytes concomitant with activation of cell sheet displacement by collective migration to restore damaged epidermal surfaces (20–23). The strong correlation between quiescence exit and cell migration in multiple physiological settings suggests the existence of mechanisms that link quiescence exit to activation of cell motility.The dynamics of epithelial collectives is largely regulated by mechanical forces generated through cell–cell interactions as well as interactions between cells and the extracellular environment (24). Key components involved in controlling these forces are cytoskeletal components such as actinomyosin and adhesion complexes such as adherent junctions and focal adhesion complexes (25). Additional factors that have been reported to influence the dynamic behavior of epithelial monolayers include the presence of epithelial edges (24, 26), mechanical stretching or compression (27, 28), expression of the endosomal Rab5 protein (29), exposure of cells to growth factors (30–32), local changes in cell shape (33), and the ability of cells to undergo neighbor exchange (34, 35). In addition, recent studies have also identified a functional link between cell cycle progression and force fluctuation leading to dynamic behavior of cultured epithelial monolayers (36, 37).In this study, we have investigated a mechanical link between quiescence exit and activation of large-scale cell sheet displacements. Using traction force microscopy (TFM), we found that confluent cell monolayers install an actinomyosin-based system during quiescence that produces a coordinated burst of contractile forces and intercellular tension across the epithelial monolayer immediately following exposure to serum-borne mitogens. By combining experiments and theoretical modeling, we show that the amplified forces are essential for driving coordinated cell sheet displacements within otherwise motility-restricted cell monolayers. Furthermore, the magnitude of mechanical forces created during quiescence exit and the extent of cell sheet displacement correlate with quiescence depth. Our study provides evidence that quiescent keratinocyte monolayers possess mechanical preparedness for motility and establish monolayer stress amplification as a strategy for overcoming the motility barrier in confined cell sheets.     

Safety evaluation of Streptomyces murinus glucose isomerase

A glucose isomerase enzyme, obtained from Streptomyces murinus, was produced by a fermentation process and subjected to a series of tests to investigate its safety in use and manufacture. It was not mutagenic (Ames test, using liquid culture) nor did it provoke chromosomal damage (rat bone marrow cytogenetics test). It did not contain (nor did the organism produce) antimicrobial activity or macrolidpolyene antibiotics. It had no teratogenic activity when administered to pregnant rats at 100,000 ppm in the diet. It was without effect upon rats when administered at this dietary concentration for 4 weeks. Dietary administration at 5000, 15,000 or 50,000 ppm to rats for 13 weeks resulted in nephrocalcinosis in females at all dosages (probably a physiological response to the altered calcium:phosphate ratio in the admixed diet) and status spongiosus in the brains of males receiving 50,000 ppm. As the finding of nephrocalcinosis in rats is generally agreed to be of no toxicological importance with regard to the use in man, the dietary concentration of 15,000 ppm was considered to be highest no-effect level. This level corresponds to an intake of some 1000 mg/kg/day, which represents approximately 8000 times the human intake based on a conservative estimation.     

Artificial Sweeteners in Breast Milk: A Clinical Investigation with a Kinetic Perspective

Artificial sweeteners (ASs) are calorie-free chemical substances used instead of sugar to sweeten foods and drinks. Pregnant women with obesity or diabetes are often recommended to substitute sugary products with ASs to prevent an increase in body weight. However, some recent controversy surrounding ASs relates to concerns about the risk of obesity caused by a variety of metabolic changes, both in the mother and the offspring. This study addressed these concerns and investigated the biodistribution of ASs in plasma and breast milk of lactating women to clarify whether ASs can transfer from mother to offspring through breast milk. We recruited 49 lactating women who were provided with a beverage containing four different ASs (acesulfame-potassium, saccharin, cyclamate, and sucralose). Blood and breast milk samples were collected before and up to six hours after consumption. The women were categorized: BMI < 25 (n = 20), BMI > 27 (n = 21) and type 1 diabetes (n = 8). We found that all four ASs were present in maternal plasma and breast milk. The time-to-peak was 30–120 min in plasma and 240–300 min in breast milk. Area under the curve (AUC) ratios in breast milk were 88.9% for acesulfame-potassium, 38.9% for saccharin, and 1.9% for cyclamate. We observed no differences in ASs distributions between the groups.     

单克隆抗体博来霉素A6偶联物对白血病细胞特异性结合与内化

抗CCT2单克隆抗体博来霉素A6偶联物可吸附胶体金颗粒(McAb-A6-Au)。电镜观察表明,在4℃,1h,表面有McAb-A6-Au颗粒的CEM细胞最高达78%;在37℃,4h,内化McAb-A6-Au颗粒的CEM细胞高达72%。而抗原性无关的U937细胞仅为14%。并且McAb-A6-Au颗粒能直接穿过细胞膜、核膜进入细胞浆和细胞核。37℃,1h已有10～18%的CEM细胞核内有McAb-A 6-Au颗粒。实验结果提示了单抗与博来霉素A6的偶联物与选择性地结合靶细胞,而且进入细胞速度快、穿透力强,有可能成为治疗白血病药物。     

High measles mortality in infancy related to intensity of exposure

In a West African urban community, measles infection in infants was examined over 5 years (1979-1983). In the age group 0 to 11 months, measles mortality was higher among secondary cases (infected in the house) than among index cases (infected outside the house), and the proportion of secondary cases was significantly higher for this age group than for older children. Intensive exposure related to the social pattern of disease transmission may be important in explaining the high infant mortality observed with measles in developing countries. Mortality during the first 12 months of life increased with age, presumably because of the decrease of maternally derived measles antibodies. In children younger than 6 months of age, who are usually considered to be protected by maternal antibody, intensive exposure may lead to infection, as demonstrated by a high level of measles-specific antibodies in some children exposed to an older sibling with measles. The aim of public health policies should be to change conditions of exposure.     

Effects of an Amino Acid-Based Formula Supplemented with Two Human Milk Oligosaccharides on Growth,Tolerability, Safety,and Gut Microbiome in Infants with Cow’s Milk Protein Allergy

This open-label, non-randomized, multicenter trial (Registration: {"type":"clinical-trial","attrs":{"text":"NCT 03661736","term_id":"NCT03661736"}}NCT 03661736) aimed to assess if an amino acid-based formula (AAF) supplemented with two human milk oligosaccharides (HMO) supports normal growth and is well tolerated in infants with a cow’s milk protein allergy (CMPA). Term infants aged 1–8 months with moderate-to-severe CMPA were enrolled. The study formula was an AAF supplemented with 2′-fucosyllactose (2′-FL) and lacto-N-neotetraose (LNnT). Infants were fed the study formula for 4 months and were offered to remain on the formula until 12 months of age. Tolerance and safety were assessed throughout the trial. Out of 32 infants (mean age 18.6 weeks; 20 (62.5%) male), 29 completed the trial. During the 4-month principal study period, the mean weight-for-age Z score (WAZ) increased from –0.31 at the baseline to +0.28 at the 4-months’ follow-up. Linear and head growth also progressed along the WHO child growth reference, with a similar small upward trend. The formula was well tolerated and had an excellent safety profile. When comparing the microbiome at the baseline to the subsequent visits, there was a significant on-treatment enrichment in HMO-utilizing bifidobacteria, which was associated with a significant increase in fecal short-chain fatty acids. In addition, we observed a significant reduction in the abundance of fecal Proteobacteria, suggesting that the HMO-supplemented study formula partially corrected the gut microbial dysbiosis in infants with CMPA.     

Assay of tolfenamic acid and its metabolites by liquid chromatography on dynamically modified silica: application in pharmacokinetics

An LC method able to separate all known metabolites of tolfenamic acid was developed. It was applied to characterize the metabolic profiles after single and multiple peroral doses of tolfenamic acid to volunteers. Up to 80% of the dose, mostly conjugated metabolites, could be recovered in the urine by this method. Two of the metabolites showed plasma half-lives of several days and peculiar conjugation properties.     

Effect of Elevation and Surface Roughness on Naturalness Perception of 2.5D Decor Prints

Naturalness is a complex concept. It can involve a variety of attributes. In this work, we considered the effect of elevation and surface roughness on naturalness perception of 2.5D decor prints for four material categories. We found that elevation has an impact on the naturalness perception of 2.5D decor prints and that it is linked with content. The observers found lower elevation to be more natural for wood and glass 2.5D prints while there was no clear tendency for stone and metal 2.5D prints. We also found the perceptual attributes used for naturalness assessment of 2.5D decor prints. The top five ones are color, roughness, gloss, elevation, and lightness. The obtained findings can be useful for companies that produce 2.5D prints.