DNA Extraction from Formalin-fixed and Paraffin-embedded Tissues by Triton X-100 for Effective Amplification of EGFR Gene by Polymerase Chain Reaction

For first-line non-small-cell lung cancer（NSCLC） therapy,detecting mutation status of the epidermal growth factor receptor（EGFR） gene constitutes a prudent test to identify patients who are most likely to benefit from EGFR-tyrosine kinase inhibitor（TKI） therapy.Now,the material for detecting EGFR gene mutation status mainly comes from formalin-fixed and paraffin-embedded（FFPE） tissues.DNA extraction from FFPE and the amplification of EGFR gene by polymerase chain reaction（PCR） are two key steps for detecting EGFR gene mutation.We showed a simple method of DNA extraction from FFPE tissues for the effective amplification of EGFR gene.Extracting DNA from the FFPE tissues of NSCLC patients with 1% Triton X-100（pH=10.0） was performed by heating at 95 °C for 30 min.Meanwhile,a commercial kit was used to extract DNA from the same FFPE tissues of NSCLC patients for comparison.DNA extracted products were used as template for amplifying the exons 18,19,20 and 21 of EGFR by PCR for different amplified fragments.Results show that DNA fragment size extracted from FFPE tissues with 1% Triton X was about 250―500 base pairs（bp）.However,DNA fragment size extracted from FFPE tissues via commercial kit was about from several hundreds to several thousands bp.The DNA yield extracted from FFPE tissues with 1% Triton X was larger than that via commercial kit.For about 500 bp fragment,four exons of EGFR could not be amplified more efficiently from extracted DNA with 1% Triton X than with commercial kit.However,for about 200 bp fragment.This simple and non-laborious protocol could successfully be used to extract DNA from FFPE tissue for the amplification of EGFR gene by PCR,further screening of EGFR gene mutation and facilitating the molecular analysis of a large number of FFPE tissues from NSCLC patients.

Engineering vascularized organotypic tissues via module assembly

Adequate vascularization is a critical determinant for the successful construction and clinical implementation of complex organotypic tissue models. Currently, low cell and vessel density and insufficient vascular maturation make vascularized organotypic tissue construction difficult,greatly limiting its use in tissue engineering and regenerative medicine. To address these limitations, recent studies have adopted pre-vascularized microtissue assembly for the rapid generation of functional tissue analogs with dense vascular networks and high cell density. In this article, we summarize the development of module assembly-based vascularized organotypic tissue construction and its application in tissue repair and regeneration, organ-scale tissue biomanufacturing, as well as advanced tissue modeling.

Selection and Validation of Reference Genes for Normalization of RT-qPCR Analysis in Developing or Abiotic-Stressed Tissues of Loquat (Eriobotrya japonica)

Loquat(Eriobotrya japonica Lindl.)is a subtropical evergreen fruit tree that produces fruits with abundant nutrients and medicinal components.Confirming suitable reference genes for a set of loquat samples before qRT-PCR experiments is essential for the accurate quantification of gene expression.In this study,eight candidate reference genes were selected from our previously published RNA-seq data,and primers for each candidate reference gene were designed and evaluated.The Cq values of the candidate reference genes were calculated by RT-qPCR in 31 different loquat samples,including 12 subgroups of developing or abiotic-stressed tissues.Different combinations of stable reference genes were screened according to a comprehensive rank,which was synthesized from the results of four algorithms,including the geNorm,NormFinder,BestKeeper andΔCt methods.The screened reference genes were verified by normalizing EjLGA1 in each subgroup.The obtained suitable combinations of reference genes for accurate normalization were GAPDH,EF1αand ACT for floral development;GAPDH,UBCE and ACT for fruit setting;EF1α,GAPDH and eIF2B for fruit ripening;ACT,EF1αand UBCE for leaves under heat stress;eIF2B,UBCE and EF1αfor leaves under freezing stress;EF1α,TUA and UBCE for leaves under salt stress;ACT,EF1αand eIF2B for immature pulp under freezing stress;ACT,UBCE and eIF2B for immature seeds under freezing stress;EF1α,eIF2B and UBCE for both immature pulp and seeds under freezing stress;UBCE,TUB and TUA for red-fleshed fruits under cold-storage stress;eIF2B,RPS3 and TUB for white-fleshed fruits under coldstorage stress;and eIF2B,UBCE and RPS3 for both red-and white-fleshed fruits under cold-storage stress.This study obtained different combinations of stable reference genes for accurate normalization in twelve subgroups of developing or abiotic-stressed tissues in loquat.To our knowledge,this is the first report to obtain stable reference genes for normalizing gene expression of abiotic-stressed tissues in E.japonica.The use of the three most stable reference genes could increase the reliability of future quantification experiments.

Biomimetic natural biomaterials for tissue engineering and regenerative medicine:new biosynthesis methods,recent advances,and emerging applications

Biomimetic materials have emerged as attractive and competitive alternatives for tissue engineering(TE)and regenerative medicine.In contrast to conventional biomaterials or synthetic materials,biomimetic scaffolds based on natural biomaterial can offer cells a broad spectrum of biochemical and biophysical cues that mimic the in vivo extracellular matrix(ECM).Additionally,such materials have mechanical adaptability,micro-structure interconnectivity,and inherent bioactivity,making them ideal for the design of living implants for specific applications in TE and regenerative medicine.This paper provides an overview for recent progress of biomimetic natural biomaterials(BNBMs),including advances in their preparation,functionality,potential applications and future challenges.We highlight recent advances in the fabrication of BNBMs and outline general strategies for functionalizing and tailoring the BNBMs with various biological and physicochemical characteristics of native ECM.Moreover,we offer an overview of recent key advances in the functionalization and applications of versatile BNBMs for TE applications.Finally,we conclude by offering our perspective on open challenges and future developments in this rapidly-evolving field.

Visceral adipose tissue predicts severity and prognosis of acute pancreatitis in obese patients

Acute pancreatitis is a common systemic inflammatory disease, manifested by a spectrum of severity, ranging from mild in the majority of patients to severe acute pancreatitis. Patients with severe acute pancreatitis suffer from severe local and systemic complications and organ failure, leading to a poor prognosis. The early recognition of the severe condition is important to improve prognosis. Obesity has risen in tandem with an increase in the severity of acute pancreatitis in recent years. Studies have revealed that adipose tissue, particularly visceral adipose tissue is associated with the prognosis of acute pancreatitis. This review discussed the role of visceral adipose tissue in obese patients with acute pancreatitis and explored the possible mechanism involved.

Detection of Low-Risk and High-Risk Oncogenic Human Papillomavirus in Archived Tissues from ENT Tumors in Burkina Faso

Human papillomavirus (HPV) is classified into high-risk HPV (HR-HPV) and HPV (LR-HPV) according to their oncogenic potential. These viruses can be found in the cervix, vagina, vulva, anus and in the ENT sphere. HPV ENT infections can lead to benign or malignant tumors in which we could find both LR-HPV and HR-HPV genotypes. The objective of this study was to investigate the genotypes of HR-HPV and LR-HPV in archived tissue samples derived from both benign and malignant tumors of the ear, nose, and throat (ENT) in Ouagadougou, Burkina Faso. One hundred and twenty formalin-fixed, paraffin-embedded archived tissues of the ENT sphere from 26 benign tumors and 94 malignant tumors were included. The tissues were first deparaffinized with xylem. The extracted DNA was used to test for high-risk and low-risk HPV by Real-Time Multiplex PCR. HPV DNA was found in 57.7% (15/26) of benign tumors and 43.61% (41/94) of malignant tumors. The prevalence of HPV infection was 46.67% (56/120) in all tumors combined. The most common HPV genotypes found were HPV 11 (34.28%), HPV 6 (30%), HPV56 (14.28%) and HPV 33 (8.57%). There were 21.43% (12/56) cases of genotypes co-infections with 10 cases of double infection and 2 cases of triple infection. Both low-risk and high-risk HPV are found in ENT tumors with relatively high HPV prevalence.

Wideband frequency-dependent dielectric properties of rat tissues exposed to low-intensity focused ultrasound in the microwave frequency range

Tissue dielectric properties can vary upon the incident of an acoustic wave.The goal of this study is to quantify this change due to the acoustoelectric effect(AE),and to obtain the frequency-dependent dielectric properties of tissues exposed to low-intensity focused ultrasound(LIFU).The dielectric properties of the blood,brain,chest muscle,heart,kidney,leg muscle,liver,lung,pancreas,and spleen of rats were measured by an open-ended coaxial probe method.The acoustic intensity of LIFU focus was 2.97 MPa(67.6 W/cm^(2)),3.95 MPa(120 W/cm^(2)),and 5.17 MPa(204 W/cm^(2)),respectively,and the measurement frequency band was 0.1–7.08 GHz.The measurement results show that with the LIFU modulation,the conductivity and dielectric constant decreased in the high-frequency band,and on the contrary,they increased in the lowfrequency band,and the larger the acoustic intensity was,the more obvious the phenomenon was.This work contributes to a better understanding of the mechanisms by which ultrasound acts on the dielectric properties of biological tissues.It is expected that the findings from this study will provide a basis that the response of tissue to LIFU modulation can be monitored by noninvasive techniques such as microwave-induced thermoacoustic imaging(MTI)and microwave imaging,present a new idea for improving the endogenous contrast between different biological tissues in MTI and acoustoelectric imaging,and possibly lead to the development of a new imaging method based on the relaxation time of tissue after LIFU modulation.

Mercury accumulation,distribution,and isotopic composition in tissues of the Collared Scops Owl(Otus lettia)

Mercury is a ubiquitous contaminant known to accumulate in wildlife,particularly bird species at higher trophic levels.Knowledge of tissue-specific Hg distributions aids our understanding of Hg bioaccumulation in organisms.In this study,one adult and three juvenile Collared Scops Owls(Otus lettia)were studied to elucidate the bioaccumulation of Hg in body tissues.Six tissues and organs(feathers,nails,heart,liver,gizzard,and muscle),as well as gastric contents,were examined for total Hg(THg)and methylmercury(MeHg)contents,Hg isotopic compositions including mass-dependent fractionation(MDF;δ202Hg)and mass-independent fractionation(MIF;Δ199Hg andΔ201Hg),and C(δ13C)and N(δ15N)isotopic compositions.Tissue-specific THg and MeHg concentrations in the adult were in the ranges of 150–1360 ng/g and17–1060 ng/g,and lower in the juveniles at 91–419 ng/g and 67–350 ng/g,respectively.Theδ^(202)Hg values in the adult were strongly negative at-1.75‰±0.17‰compared with the juveniles at-0.99‰±0.25‰.The adult exhibited lower MIF values than the juveniles,at0.23‰±0.07‰forΔ^(199)Hg and 0.2‰±0.11‰forΔ^(201)Hg,comparedwith0.81‰±0.09‰and0.66‰±0.07‰,respectively.The lower adult MDF and MIF values suggest that the adult tended to accumulate negative Hg isotopes but the juvenile's positive Hg isotopes.Differences between adult and juvenile tissue Hg concentrations indicate that metabolic processes play an important role in Hg accumulation.

Heat transport properties within living biological tissues with temperature-dependent thermal properties

Understanding of the heat transport within living biological tissues is crucial to effective heat treatments. The heat transport properties of living biological tissues with temperature-dependent properties are explored in this paper. Taking into account of variable physical properties, the governing equation of temperature is first derived in the context of the dualphase-lags model(DPL). An effective method, according to the Laplace transform and a linearization technique, is then employed to solve this nonlinear governing equation. The temperature distribution of a biological tissue exposed to a pulsed heat flux on its exterior boundary, which frequently happens in various heat treatments, is predicted and analyzed. The results state that a lower temperature can be predicted when temperature dependence is considered in the heating process.The contributions of key thermal parameters are different and dependent on the ratio of phase lag and the amplitude of the exterior pulsed heat flux.

Biomaterials and tissue engineering in traumatic brain injury:novel perspectives on promoting neural regeneration

Traumatic brain injury is a serious medical condition that can be attributed to falls, motor vehicle accidents, sports injuries and acts of violence, causing a series of neural injuries and neuropsychiatric symptoms. However, limited accessibility to the injury sites, complicated histological and anatomical structure, intricate cellular and extracellular milieu, lack of regenerative capacity in the native cells, vast variety of damage routes, and the insufficient time available for treatment have restricted the widespread application of several therapeutic methods in cases of central nervous system injury. Tissue engineering and regenerative medicine have emerged as innovative approaches in the field of nerve regeneration. By combining biomaterials, stem cells, and growth factors, these approaches have provided a platform for developing effective treatments for neural injuries, which can offer the potential to restore neural function, improve patient outcomes, and reduce the need for drugs and invasive surgical procedures. Biomaterials have shown advantages in promoting neural development, inhibiting glial scar formation, and providing a suitable biomimetic neural microenvironment, which makes their application promising in the field of neural regeneration. For instance, bioactive scaffolds loaded with stem cells can provide a biocompatible and biodegradable milieu. Furthermore, stem cells-derived exosomes combine the advantages of stem cells, avoid the risk of immune rejection, cooperate with biomaterials to enhance their biological functions, and exert stable functions, thereby inducing angiogenesis and neural regeneration in patients with traumatic brain injury and promoting the recovery of brain function. Unfortunately, biomaterials have shown positive effects in the laboratory, but when similar materials are used in clinical studies of human central nervous system regeneration, their efficacy is unsatisfactory. Here, we review the characteristics and properties of various bioactive materials, followed by the introduction of applications based on biochemistry and cell molecules, and discuss the emerging role of biomaterials in promoting neural regeneration. Further, we summarize the adaptive biomaterials infused with exosomes produced from stem cells and stem cells themselves for the treatment of traumatic brain injury. Finally, we present the main limitations of biomaterials for the treatment of traumatic brain injury and offer insights into their future potential.

Three-dimensional remodeling of collagen fibers within cervical tissues in pregnancy

The cervix is a collagen-rich connective tissue that must remain closed during pregnancy while undergoing progressive remodeling in preparation for delivery,which begins before the onset of the preterm labor process.Therefore,it is important to resolve the changes of collagen flbers during cervical remodeling for the prevention of preterm labor.Herein,we assessed the spatial organization of collagen flbers in a three-dimensional(3D)context within cervical tissues of mice on day 3,9,12,15 and 18 of gestation.We found that the 3D directional variance,a novel metric of alignment,was higher on day 9 than that on day 3 and then gradually decreased from day 9 to day 18.Compared with two-dimensional(2D)approach,a higher sensitivity was achieved from 3D analysis,highlighting the importance of truly 3D quantification.Moreover,the depthdependent variation of 3D directional variance was investigated.By combining multiple 3D directional variance-derived metrics,a high level of classification accuracy was acquired in distinguishing different periods of pregnancy.These results demonstrate that 3D directional variance is sensitive to remodeling of collagen fibers within cervical tissues,shedding new light on highly-sensitive,early detection of preterm birth(PTB).

Depolarizing metrics in the biomedical field:Vision enhancement and classification of biological tissues

Polarimetry encompasses a collection of optical techniques broadly used in a variety of fields.Nowadays,such techniques have provided their suitability in the biomedical field through the study of the polarimetric response of biological samples(retardance,dichroism and depolarization)by measuring certain polarimetric observables.One of these features,depolarization,is mainly produced by scattering on samples,which is a predominant efiect in turbid media as biological tissues.In turn,retardance and dichroic efiects are produced by tissue anisotropies and can lead to depolarization too.Since depolarization is a predominant efiect in tissue samples,we focus on studying difierent depolarization metrics for biomedical applications.We report the suitability of a set of depolarizing observables,the indices of polarimetric purity(IPPs),for biological tissue inspection.We review some results where we demonstrate that IPPs lead to better performance than the depolarization index,which is a well-established and commonly used depolarization observable in the literature.We also provide how IPPs are able to significantly enhance contrast between difierent tissue structures and even to reveal structures hidden by using standard intensity images.Finally,we also explore the classificatory potential of IPPs and other depolarizing observables for the discrimination of difierent tissues obtained from ex vivo chicken samples(muscle,tendon,myotendinous junction and bone),reaching accurate models for tissue classification.

Effects of adjacent bubble on spatiotemporal evolutions of mechanical stresses surrounding bubbles oscillating in tissues

The cavitation dynamics and mechanical stress in viscoelastic tissues, as the primary mechanisms of some ultrasound therapies, are extremely complex due to the interactions of cavitation bubble with adjacent bubbles and surrounding tissues.Therefore, the cavitation dynamics and resultant mechanical stress of two-interacting bubbles in the viscoelastic tissues are numerically investigated, especially focusing on the effects of the adjacent bubble. The results demonstrate that the mechanical stress is highly dependent on the bubble dynamics. The compressive stress and tensile stress are generated at the stage of bubble expansion and collapse stage, respectively. Furthermore, within the initial parameters examined in this paper, the effects of the adjacent bubble will distinctly suppress the radial expansion of the small bubble and consequently lead its associated stresses to decrease. Owing to the superimposition of two stress fields, the mechanical stresses surrounding the small bubble in the direction of the neighboring bubble are smaller than those in other directions. For two interacting cavitation bubbles, the suppression effects of the nearby bubble on both the cavitation dynamics and the stresses surrounding the small bubble increase as the ultrasound amplitude and the initial radius of the large bubble increase, whereas they decrease with the inter-bubble distance increasing. Moreover, increasing the tissue viscoelasticity will reduce the suppression effects of the nearby bubble, except in instances where the compressive stress and tensile stress first increase and then decrease with the tissue elasticity and viscosity increasing respectively. This study can provide a further understanding of the mechanisms of cavitation-associated mechanical damage to the adjacent tissues or cells.

Interplay between mesenchymal stem cells and macrophages:Promoting bone tissue repair

The repair of bone tissue damage is a complex process that is well-orchestrated in time and space,a focus and difficulty in orthopedic treatment.In recent years,the success of mesenchymal stem cells(MSCs)-mediated bone repair in clinical trials of large-area bone defects and bone necrosis has made it a candidate in bone tissue repair engineering and regenerative medicine.MSCs are closely related to macrophages.On one hand,MSCs regulate the immune regulatory function by influencing macrophages proliferation,infiltration,and phenotype polarization,while also affecting the osteoclasts differentiation of macrophages.On the other hand,macrophages activate MSCs and mediate the multilineage differentiation of MSCs by regulating the immune microenvironment.The cross-talk between MSCs and macrophages plays a crucial role in regulating the immune system and in promoting tissue regeneration.Making full use of the relationship between MSCs and macrophages will enhance the efficacy of MSCs therapy in bone tissue repair,and will also provide a reference for further application of MSCs in other diseases.

Characteristics and mechanisms of subcutaneous and visceral adipose tissue aging

Aging is one of the most significant health challenges worldwide and is a primary cause of chronic diseases and physiological decline.Among the myriad changes that occur with aging,alterations in adipose tissue distribution and function have gained considerable attention because of their profound impact on metabolic health and overall well-being.Subcutaneous adipose tissue(SAT)and visceral adipose tissue(VAT)are the two major depots of white adipose tissue,each with distinct roles in metabolism and health.Understanding the characteristics and underlying mechanisms of SAT and VAT is crucial for elucidating the aging process and developing strategies to promote healthy aging.This review focuses on delineating and analyzing the characteristics and intrinsic mechanisms underlying the aging of subcutaneous and visceral adipose tissue during the aging process,which can contribute to a better understanding of the aging process and enhance healthy aging.

Research on the Development of Fibroin and Nano-Fiber from Silk Cocoons for Regenerated Tissue Engineering Applications by Electro-Spinning

In this paper, the main goal is to prepare silk fibroin nano-fiber, which is used for regenerated tissue applications. Silk scaffold nano-fibers made by electro-spinning technology can be used in regenerated tissue applications. The purpose of the research is to prepare a silk-fibroin nano-fiber solution for potential applications in tissue engineering. Using a degumming process, pure silk fibroin protein is extracted from silk cocoons. The protein solution for fibroin is purified, and the protein content is determined. The precise chemical composition, exact temperature, time, voltage, distance, ratio, and humidity all have a huge impact on degumming, solubility, and electro-spinning nano-fibers. The SEM investigates the morphology of silk fibroin nano-fibres at different magnifications. It also reveals the surface condition, fiber orientation, and fiber thickness of the silk fibroin nano-fiber. The results show that regenerated silk fibroin and nano-fiber can be used in silk fibroin scaffolds for various tissue engineering applications.

Magnesium-incorporated biocomposite scaffolds:A novel frontier in bone tissue engineering

Nonunion represents a crucial challenge in orthopedic medicine,demanding innovative solutions beyond the scope of traditional bone grafting methods.Among the various strategies available,magnesium(Mg)implants have been recognized for their biocompatibility and biodegradability.However,their susceptibility to rapid corrosion and degradation has garnered notable research interest in bone tissue engineering(BTE),particularly in the development of Mg-incorporated biocomposite scaffolds.These scaffolds gradually release Mg2+,which enhances immunomodulation,osteogenesis,and angiogenesis,thus facilitating effective bone regeneration.This review presents myriad fabrication techniques used to create Mg-incorporated biocomposite scaffolds,including electrospinning,three-dimensional printing,and sol-gel synthesis.Despite these advancements,the application of Mg-incorporated biocomposite scaffolds faces challenges such as controlling the degradation rate of Mg and ensuring mechanical stability.These limitations highlight the necessity for ongoing research aimed at refining fabrication techniques to better regulate the physicochemical and osteogenic properties of scaffolds.This review provides insights into the potential of Mg-incorporated biocomposite scaffolds for BTE and the challenges that need to be addressed for their successful translation into clinical applications.

Engineering of ovarian tissue for ovarian dysfunctions:A review

This review explores tissue engineering as a potential solution for reproductive health issues in women caused by genetic or acquired diseases,such as premature ovarian failure or oophorectomy.The loss of ovarian function can lead to infertility,osteoporosis,and cardiovascular disease.Hormone replacement therapy is a common treatment,but it has limitations and risks.The review focuses on two main approaches in tissue engineering:scaffold-based(3D printing,electrospinning,decellularization)and scaffold-free(stem cell transplantation,organoid cultivation).Both approaches show promise in preclinical studies for creating functional ovarian tissue.Challenges include vascularization,innervation,long-term function,and safety.Despite these challenges,tissue engineering offers a potential avenue for restoring fertility and hormone balance in women with ovarian dysfunction.

A Review on Silk Fibroin as a Biomaterial in Tissue Engineering

Regenerative medicine progress is based on the development of cell and tissue bioengineering. One of the aims of tissue engineering is the development of scaffolds, which should substitute the functions of the replaced organ after their implantation into the body. The tissue engineering material must meet a range of requirements, including biocompatibility, mechanical strength, and elasticity. Furthermore, the materials have to be attractive for cell growth: stimulate cell adhesion, migration, proliferation and differentiation. One of the natural biomaterials is silk and its component (silk fibroin). An increasing number of scientists in the world are studying silk and silk fibroin. The purpose of this review article is to provide information about the properties of natural silk (silk fibroin), as well as its manufacture and clinical application of each configuration of silk fibroin in medicine. Materials and research methods. Actual publications of foreign authors on resources PubMed, Medline, E-library have been analyzed. The selection criteria were materials containing information about the structure and components of silk, methods of its production in nature. This article placed strong emphasis on silk fibroin, the ways of artificial modification of it for use in various sphere of medicine.

Endotoxin-induced alterations of adipose tissue function:a pathway to bovine metabolic stress

During the periparturient period, dairy cows exhibit negative energy balance due to limited appetite and increased energy requirements for lactogenesis. The delicate equilibrium between energy availability and expenditure puts cows in a state of metabolic stress characterized by excessive lipolysis in white adipose tissues(AT), increased production of reactive oxygen species, and immune cell dysfunction. Metabolic stress, especially in AT, increases the risk for metabolic and inflammatory diseases. Around parturition, cows are also susceptible to endotoxemia. Bacterial-derived toxins cause endotoxemia by promoting inflammatory processes and immune cell infiltration in different organs and systems while impacting metabolic function by altering lipolysis, mitochondrial activity, and insulin sensitivity. In dairy cows, endotoxins enter the bloodstream after overcoming the defense mechanisms of the epithelial barriers, particularly during common periparturient conditions such as mastitis, metritis, and pneumonia, or after abrupt changes in the gut microbiome. In the bovine AT, endotoxins induce a pro-inflammatory response and stimulate lipolysis in AT, leading to the release of free fatty acids into the bloodstream. When excessive and protracted, endotoxin-induced lipolysis can impair adipocyte's insulin signaling pathways and lipid synthesis. Endotoxin exposure can also induce oxidative stress in AT through the production of reactive oxygen species by inflammatory cells and other cellular components. This review provides insights into endotoxins' impact on AT function, highlighting the gaps in our knowledge of the mechanisms underlying AT dysfunction, its connection with periparturient cows' disease risk, and the need to develop effective interventions to prevent and treat endotoxemia-related inflammatory conditions in dairy cattle.