A simple and fast alternative method to remove glycerol from chicken semen after cryopreservation.

A concentration of 11% of glycerol is the standard one for sperm cryopreservation in chickens, however, the presence of just 2% glycerol already causes severe fertility reduction, suggesting the necessity of removing glycerol before artificial insemination (AI). The major approach developed for this purpose is serial dilution followed by centrifugation (SDC), which demands special equipment (such as a refrigerate room) to maintain post-thaw semen at 4 °C, besides being time consuming. Therefore, we attempted to develop a simple method to remove glycerol from chicken frozen-thawed semen based on a colloidal gel, Percoll, which is ordinarily used to select motile and viable sperm in mammals as well as in fresh chicken semen. In this study, we used a Percoll based glycerol removal solution (GRS) containing sucrose to avoid frozen-thawed sperm suffering from osmotic stress. Subsequently, several conditions including GRS compositions (GRS A, B, C and D) and centrifugation temperatures (4 and 20 °C) were compared by their influence on sperm in vitro parameters. Afterwards, GRS A and D were selected for fertility evaluation, compared to conventional SDC method. Our results showed that the fertility with GRS A at both 4 and 20 °C were higher than GRS D (p < 0.05) and similar or even superior to the fertility obtained with SDC method. Altogether, our novel GRS protocol is a valuable method for chicken sperm cryobanking policy, supported by its notable results of fertility as well as saving 44% of time, with a simple equipment at flexible operation temperatures of 4 or 20 °C.

Dopaminergic neurons derived from porcine induced pluripotent stem cell like cells function in the Lanyu pig model of Parkinson's disease.

The induced pluripotent stem cells (iPSCs) are able to differentiate into dopaminergic neurons and execute the therapeutic effects for Parkinson's disease (PD). Here, we established a animal model of PD in Lanyu pigs by injecting 5 mg/kg of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP). Next, the porcine iPSC-like cells (piPSC-like cells) were differentiated into D18 neuronal progenitors (D18 NPs) that were transplanted into the striatum to evaluate their therapeutic effects of PD. We showed that after 8 weeks of cell transplantation, the behavior score was significantly ameliorated and fully recovered at the 14th week of cell transplantation. The number of dopaminergic neurons was also significantly improved at the end of the experiment although the number was still about 50% lower than that in the control group. Our findings suggest that piPSC-like cell-derived D18 NPs exhibit a potential for the treatment of PD in the Lanyu pig model.

Improvement of output efficiency of p-face up photonic-crystal surface-emitting lasers.

We optimized the p-side emission device configuration of photonic-crystal surface-emitting laser (PCSEL) to facilitate the easier chip process and wafer level testing as well as the feasibility of lasing at shorter wavelength. Typically, in order to obtain uniformly distributed current for larger emission area of PCSELs, laser output is designed through the n-side window due to the low hole mobility and thin p-side cladding layer. However, the substrate as well as the epi-layers have to be isolated before the test of each single die on the wafer, which compromised the advantage of wafer-level test of surface emitters. On the other hand, for lasers with emission photon energy higher than the bandgap energy of GaAs substrate, the power will be entirely attenuated. In this study, the optimized p-side emission by applying the transparent conduction layer on top of the p side contact layer to enhance the current distribution and breaking the symmetry of conventional circle pattern in a unit cell to boost the output efficiency is investigated. Through this approach, a high efficiency p-side up PCSEL platform with lower fabrication cost is developed, which is also applicable for short wavelength PCSELs.

Inhibition of the formation of autophagosome but not autolysosome augments ABT-751-induced apoptosis in TP53-deficient Hep-3B cells.

The objective was to investigate the upstream mechanisms of apoptosis which were triggered by a novel antimicrotubule drug, ABT-751, in a tumor protein p53 ( TP53)-deficient hepatocellular carcinoma-derived Hep-3B cells. A series of in vitro assays indicated that ABT-751 caused the disruption of the mitotic spindle structure, collapse of mitochondrial membrane potential, generation of reactive oxygen species, DNA damage, G 2 /M cell cycle arrest, inhibition of anchorage-independent cell growth and apoptosis in Hep-3B cells accompanied by alteration of the expression levels of several DNA damage checkpoint proteins and cell cycle regulators. Subsequently, ABT-751 triggered apoptosis along with markedly upregulated several proapoptotic proteins involving in extrinsic, intrinsic, and caspase-mediated apoptotic pathways. A pan-caspase inhibitor suppressed ABT-751-induced apoptosis. ABT-751 also induced autophagy soon after the occurrence of apoptosis through the suppression of AKT serine/threonine kinase/mechanistic target of rapamycin signaling pathway. Exogenous expression of the TP53 gene significantly incurred both apoptosis and autophagy in Hep-3B cells. Pharmacological inhibition of autophagosome (early autophagy) but not autolysosome (late autophagy) enhanced ABT-751-induced apoptosis in TP53-deficient Hep-3B cells. Our study provided a new strategy to augment ABT-751-induced apoptosis in TP53-deficient cells.

Pathogenic Acanthamoeba castellanii Secretes the Extracellular Aminopeptidase M20/M25/M40 Family Protein to Target Cells for Phagocytosis by Disruption.

Acanthamoeba is free-living protist pathogen capable of causing a blinding keratitis and granulomatous encephalitis. However, the mechanisms of Acanthamoeba pathogenesis are still not clear. Here, our results show that cells co-cultured with pathogenic Acanthamoeba would be spherical and floated, even without contacting the protists. Then, the Acanthamoeba protists would contact and engulf these cells. In order to clarify the contact-independent pathogenesis mechanism in Acanthamoeba, we collected the Acanthamoeba-secreted proteins (Asp) to incubate with cells for identifying the extracellular virulent factors and investigating the cytotoxicity process. The Asps of pathogenic Acanthamoeba express protease activity to reactive Leu amino acid in ECM and induce cell-losing adhesion ability. The M20/M25/M40 superfamily aminopeptidase protein (ACA1_264610), an aminopeptidase be found in Asp, is upregulated after Acanthamoeba and C6 cell co-culturing for 6 h. Pre-treating the Asp with leucine aminopeptidase inhibitor and the specific antibodies of Acanthamoeba M20/M25/M40 superfamily aminopeptidase could reduce the cell damage during Asp and cell co-incubation. These results suggest an important functional role of the Acanthamoeba secreted extracellular aminopeptidases in the Acanthamoeba pathogenesis process. This study provides information regarding clinically pathogenic isolates to target specific molecules and design combined drugs.

A microtubule inhibitor, ABT-751, induces autophagy and delays apoptosis in Huh-7 cells.

The objective was to investigate the upstream mechanisms of apoptosis which were triggered by a novel anti-microtubule drug, ABT-751, in hepatocellular carcinoma-derived Huh-7 cells. Effects of ABT-751 were evaluated by immunocytochemistry, flow cytometric, alkaline comet, soft agar, immunoblotting, CytoID, green fluorescent protein-microtubule associated protein 1 light chain 3 beta detection, plasmid transfection, nuclear/cytosol fractionation, coimmunoprecipitation, quantitative reverse transcription-polymerase chain reaction, small-hairpin RNA interference and mitochondria/cytosol fractionation assays. Results showed that ABT-751 caused dysregulation of microtubule, collapse of mitochondrial membrane potential, generation of reactive oxygen species (ROS), DNA damage, G2/M cell cycle arrest, inhibition of anchorage-independent cell growth and apoptosis in Huh-7 cells. ABT-751 also induced early autophagy via upregulation of nuclear TP53 and downregulation of the AKT serine/threonine kinase (AKT)/mechanistic target of rapamycin (MTOR) pathway. Through modulation of the expression levels of DNA damage checkpoint proteins and G2/M cell cycle regulators, ABT-751 induced G2/M cell cycle arrest. Subsequently, ABT-751 triggered apoptosis with marked downregulation of B-cell CLL/lymphoma 2, upregulation of mitochondrial BCL2 antagonist/killer 1 and BCL2 like 11 protein levels, and cleavages of caspase 8 (CASP8), CASP9, CASP3 and DNA fragmentation factor subunit alpha proteins. Suppression of ROS significantly decreased ABT-751-induced autophagic and apoptotic cells. Pharmacological inhibition of autophagy significantly increased the percentages of ABT-751-induced apoptotic cells. The autophagy induced by ABT-751 plays a protective role to postpone apoptosis by exerting adaptive responses following microtubule damage, ROS and/or impaired mitochondria.

Evaluation of cellular retinoic acid binding protein 2 gene expression through the retinoic acid pathway by co-incubation of Blastocystis ST-1 with HT29 cells in vitro.

Blastocystis is a parasitic protist with a worldwide distribution that is commonly found in patients with colon and gastrointestinal pathological symptoms. Blastocystis infection has also commonly been reported in colorectal cancer and HIV/AIDS patients with gastrointestinal symptoms. To understand the pathway networks of gene regulation and the probable mechanisms influencing functions of HT-29 host cells in response to parasite infection, we examined the expression of 163 human oncogenes and kinases in human colon adenocarcinoma HT-29 cells co-incubated with Blastocystis by in-house cDNA microarray and PCR analysis. At least 10 genes were shown to be modified following Blastocystis co-incubation, including those with immunological, tumorigenesis, and antitumorigenesis functions. The expression of genes encoding cellular retinoic acid binding protein 2 (CRABP2) and proliferating cell nuclear antigen (PCNA) was markedly upregulated and downregulated, respectively. Reverse transcriptase-PCR validated the modified transcript expression of CRABP2 and other associated genes such as retinoic acid (RA)-related nuclear-receptor (RARα). Together, our data indicate that CRABP2, RARα, and PCNA expressions are involved in RA signaling regulatory networks that affect the growth, proliferation, and inflammation of HT-29 cells.

Clinical aggressiveness of myxofibrosarcomas associates with down-regulation of p12CDK2AP1: prognostic implication of a putative tumor suppressor that induces cell cycle arrest and apoptosis via mitochondrial pathway.

BACKGROUND: Attenuated endogenous protein levels of cyclin-dependent kinase 2 associated protein 1 (p12(CDK2AP1)) and its active homodimer p25(CDK2AP1) were found in myxofibrosarcoma-derived cell lines. Clinical and biological significances of this putative tumor suppressor in myxofibrosarcoma were studied. METHODS: Plasmids carrying the CDK2AP1 gene and small hairpin RNA interference (shRNAi) targeting CDK2AP1 were transfected into NMFH-1 and/or OH931 cells to evaluate the effects on the CDK2, active caspase 3 (CASP3), cleaved-CASP8 and -CASP9 levels, cell cycle regulation, and/or apoptotic responses. Immunostaining of p12(CDK2AP1) was interpretable in 102 primary myxofibrosarcomas and correlated with clinicopathological variables, CDK2, Ki-67 and active CASP3 protein levels, and disease-specific survival. RESULTS: Exogenous expression of p12(CDK2AP1) in NMFH-1 and OH931 cells significantly induced G0/G1 cell cycle arrest and down-regulated CDK2 protein level. In NMFH-1 cells, these aspects were reversed by shRNAi targeting CDK2AP1 gene. Increased active CASP3 and cleaved-CASP9, but not -CASP8, were detected after CDK2AP1 overexpression, suggesting the cellular apoptosis were induced through the mitochondrial pathway. Immunostains of p12(CDK2AP1) were aberrantly decreased in 56.9 % of cases; positively and negatively correlated with protein levels of CDK2 (p = 0.023), Ki-67 (p = 0.001) and active CASP3 (p < 0.001), respectively. Following by high histological grades, p12(CDK2AP1) down-regulation was predictive of worse disease-specific survival in univariate (p = 0.003) and multivariate (p = 0.004) analyses. CONCLUSIONS: Through down-regulation of CDK2, high p12(CDK2AP1) level induced cell cycle arrest and the mitochondrial-dependent apoptotic pathway. Low p12(CDK2AP1) level represents a poor prognostic factor in patients with myxofibrosarcoma.

The E2F transcription factor 1 transactives stathmin 1 in hepatocellular carcinoma.

BACKGROUND: Through data mining the Stanford Microarray Database, the stathmin 1 (STMN1) transcript was found to be frequently upregulated in the hepatocellular carcinoma (HCC) with low alpha-fetoprotein level. The molecular mechanism of STMN1 upregulation in HCCs remained unclear. METHODS: Quantitative RT-PCR, immunoblotting, immunohistochemistry, and transfection of expression or small hairpin RNA interference plasmids, chromatin immunoprecipitation (ChIP), and quantitative ChIP assays were performed in HCC specimens or 2 distinct HCC-derived cell lines. Dual luciferase assay and site-directed mutagenesis were applied to analyze the activities of STMN1 proximal promoter region. RESULTS: STMN1 mRNA and proteins were significantly associated with several clinicopathological features. High STMN1 or E2F1 immunoexpression was predictive of poor overall survival (OS) rate (P < .01). In HCC-derived cell lines, E2F1 was elevated before STMN1 mRNA during the cell cycle. Exogenous expression of E2F1 or both transcription factor DP-1 (TFDP1) and E2F1 genes induced E2F1 and STMN1 mRNA (P < .01). Knockdown of the E2F1 gene suppressed E2F1 and STMN1 mRNA and E2F1 and STMN1 protein levels (P < .05). The promoter activity of STMN1 gene increased with overexpression of both E2F1 and TFDP1 genes (P < .05); however, it decreased when mutations were introduced in the E2F1-binding sites (P < .05). CONCLUSIONS: Upregulation of E2F1 and STMN1 proteins associate with worse outcomes in patients with HCC. E2F1 significantly correlates with STMN1 protein level in HCC lesions and in vitro transactivation assays, suggesting that STMN1 gene is transactivated by the E2F1 protein.

Coarctation-induced degenerative abdominal aortic aneurysm in a porcine model.

OBJECTIVE: Hemodynamic stress participates in the initiation and progression of aneurysmal degeneration. Coarctation increases flow-mediated stress on the aortic wall. We tested the hypothesis that prolonged coarctation of an infrarenal abdominal aorta (AA) segment leads to abdominal aortic aneurysm (AAA) formation in mini pigs. METHODS: An asymmetric, funnel-shaped flow path was created by constricting the infrarenal AA segment of Taiwanese Lanyu mini pigs (age, 7-10 months; male and female) wrapped with an 8-mm-wide expanded polytetrafluoroethylene Teflon strip for 4 weeks (4w), 8 weeks (8w), and 12 weeks (12w) (seven pigs per group). This mimics the tortuous aneurysm neck in human AAA, which increases downstream flow-mediated stress. Significant flow disturbance resulting from moderate coarctation was indicated by a pulsatility index reduced to one third the inherent levels. Sham control pigs received Teflon wrapping without coarctation. RESULTS: Aneurysm characterized by progressive medial degeneration occurred at the terminal AA after 12w coarctation. The outer dimension enlargement of the distal AA exceeded 50% compared with that of the proximal AA at 4w, 8w, and 12w postcoarctation (sham, 1.0; 4w, 1.7 ± 0.08; 8w, 1.5 ± 0.09; 12w, 1.7 ± 0.01). Lumen ratio of the distal-to-suprarenal AA increased time dependently, with 12w postcoarctation exhibiting significant increase (sham, 1.0 ± 0.05; 4w, 1.1 ± 0.11; 8w, 1.4 ± 0.20; 12w, 1.5 ± 0.09). In the distal AA, elastic lamellae exhibited fragmentation at 4w and more pronounced fragmentation with decreased density at 8w and 12w postcoarctation. Medial collagen density exhibited the trend to increase at 4w and 8w but was reversed at 12w postcoarctation. Smooth muscle exhibited disarray and nuclear density decrease at 8w and 12w postcoarctation (sham, 6966 ± 888/mm; 4w, 5747 ± 1340/mm; 8w, 4153 ± 323/mm; 12w, 4083 ± 465/mm). Gelatin zymography revealed that matrix metalloproteinase-9 activity markedly increased at 4w postcoarctation. CONCLUSIONS: Prolonged moderate coarctation caused regional hemodynamic stress and thereby induced degenerative AAA in the terminal AA.

Transplantation of porcine embryonic stem cells and their derived neuronal progenitors in a spinal cord injury rat model.

BACKGROUND AIMS: The purpose of this study was to investigate therapeutic potential of green fluorescent protein expressing porcine embryonic stem (pES/GFP(+)) cells in A rat model of spinal cord injury (SCI). METHODS: Undifferentiated pES/GFP(+) cells and their neuronal differentiation derivatives were transplanted into the contused spinal cord of the Long Evans rat, and in situ development of the cells was determined by using a live animal fluorescence optical imaging system every 15 days. After pES/GFP(+) cell transplantation, the behavior functional recovery of the SCI rats was assessed with the Basso, Beattie, and Bresnahan Locomotor Rating Scale (BBB scale), and the growth and differentiation of the grafted pES/GFP(+) cells in the SCI rats were analyzed by immunohistochemical staining. RESULTS: The relative green fluorescent protein expression level was decreased for 3 months after transplantation. The pES/GFP(+)-derived cells positively stained with neural specific antibodies of anti-NFL, anti-MBP, anti-SYP and anti-Tuj 1 were detected at the transplanted position. The SCI rats grafted with the D18 neuronal progenitors showed a significant functional recovery of hindlimbs and exhibited the highest BBB scale score of 15.20 ± 1.43 at week 24. The SCI rats treated with pES/GFP(+)-derived neural progenitors demonstrated a better functional recovery. CONCLUSIONS: Transplantation of porcine embryonic stem (pES)-derived D18 neuronal progenitors has treatment potential for SCI, and functional behavior improvement of grafted pES-derived cells in SCI model rats suggests the potential for further application of pES cells in the study of replacement medicine and functionally degenerative pathologies.

Transplantation of embryonic stem cells improves the regeneration of periodontal furcation defects in a porcine model.

OBJECTIVES: Stem cell-based therapy promises to regenerate lost tissue. Embryonic stem (ES) cells are pluripotent and may provide a virtually unlimited source for transplantation. We investigated whether ES cell transplantation improved the regeneration of furcation defects in a porcine model. MATERIAL AND METHODS: Experimental periodontitis was induced in the buccal furcations of the bilateral mandibular 2nd premolars of six minipigs. After 4 weeks, the lesions were surgically debrided and implanted with collagen matrix alone (control site) or collagen matrix overlaid with porcine ES cells expressing green fluorescent protein (pES/GFP(+) ) (test site). After 3 months of healing, the clinical parameters were measured again. The treated teeth with adjacent tissue, and part of the major organs, were processed for GFP immunohistochemistry. RESULTS: We found no obvious teratoma or rejection. The test group had significantly better clinical parameters. Immunohistochemistry (IHC) showed that transplanted pES/GFP(+) cells had differentiated to new periodontal ligament and cementum in the test sites. Surprisingly, GFP(+) cells were also detectable in the repaired control cementum and remote organs. CONCLUSIONS: We conclude that using ES cells to improve the regeneration of periodontal furcation defects is feasible. More studies are required to assess this potential treatment's efficacy and safety.

Design of low-threshold photonic-crystal surface-emitting lasers with confined gain regions by using selective area intermixing.

Photonic-crystal surface-emitting lasers have many promising properties over traditional semiconductor lasers and are regarded as the next-generation laser sources. However, the minimum achievable lasing threshold of PCSELs is still several times larger than that of VCSELs, and limiting its applications especially if the required power is small. Here, we propose a new design that reduces the gain region in the lateral plane by using selective quantum-well intermixing to reduce the threshold current of PCSELs. By performing theoretical calculations, we confirmed that the threshold current can be lowered by a factor of two to three while keeping the PCSEL's advantage of small divergence angle.

Is glycerol a good cryoprotectant for sperm cells? New exploration of its toxicity using avian model.

Glycerol is a cryoprotectant used widely for the cryopreservation of animal sperm, but it is linked to a decrease in fertility. The mechanism underlying the negative effects of glycerol remains unclear. Therefore, in this study, we aimed to gain a better understanding by using the chicken model. First, we investigated the impact of increasing the concentration of glycerol during insemination on hen fertility. Our findings revealed that 2% glycerol resulted in partial infertility, while 6% glycerol led to complete infertility. Subsequently, we examined the ability of sperm to colonize sperm storage tubules (SST) during in vivo insemination and in vitro incubation. The sperm used in the experiment were stained with Hoechst and contained 0, 2, or 6% glycerol. Furthermore, we conducted perivitelline membrane lysis tests and investigated sperm motility, mitochondrial function, ATP concentration, membrane integrity, and apoptosis after 60 min of incubation with different glycerol concentrations (0%, 1%, 2%, 6%, and 11%) at two temperatures to simulate pre-freezing (4 °C) and post-insemination (41 °C) conditions. Whereas 2% glycerol significantly reduced 50% of sperm containing SST, 6% glycerol completely inhibited SST colonization in vivo. On the other hand, in vitro incubation of sperm with SST revealed no effect of 2% glycerol, and 6% glycerol showed only a 17% reduction in sperm-filled SST. Moreover, glycerol reduced sperm-egg penetration rates and also affected sperm motility, bioenergetic metabolism, and cell death at 4 °C. These effects were observed when the concentration of glycerol exceeded 6%. Furthermore, at 41 °C, glycerol caused even greater damage, particularly in terms of reducing sperm motility. These data altogether reveal important effects of glycerol on sperm biology, sperm migration, SST colonization, and oocyte penetration. This suggests that glycerol plays a role in reducing fertility and presents opportunities for improving sperm cryopreservation.

The autocrine glycosylated-GREM1 interacts with TGFB1 to suppress TGFß/BMP/SMAD-mediated EMT partially by inhibiting MYL9 transactivation in urinary carcinoma.

PURPOSE: Urothelial carcinoma (UC) is a common disease in developed counties. This study aimed to identify autocrine roles and signaling pathways of gremlin 1, DAN family BMP antagonist (GREM1), which inhibits tumor growth and epithelial-mesenchymal transition (EMT) in UC. METHODS: Systematic in vitro and in vivo studies using genetic engineering, different urinary bladder urothelial carcinoma (UBUC)-derived cell lines, and mouse models were performed, respectively. Further, primary upper tract urothelial carcinoma (UTUC) and UBUC specimens were evaluated by immunohistochemistry. RESULTS: GREM1 protein levels conferred better disease-specific and metastasis-free survival rates and played an independent prognostic factor in UTUC and UBUC. Hypermethylation is the primary cause of low GREM1 levels. In different UBUC-derived cell lines, the autocrine/secreted and glycosylated GREM1 interacted with transforming growth factor beta 1 (TGFB1) and inhibited TGFß/BMP/SMAD signaling and myosin light chain 9 (MYL9) transactivation, subsequently cell proliferation and epithelial-mesenchymal transition (EMT). Secreted and glycosylated GREM1 also suppressed tumor growth, metastasis, and MYL9 levels in the mouse model. Instead, cytosolic GREM1 promoted cell proliferation and EMT by activating the tumor necrosis factor (TNF)/AKT/nuclear factor kappa B (NFκB) axis. CONCLUSIONS: Clinical associations, animal models, and in vitro indications provided solid evidence to show that the epithelial autocrine GREM1 is a novel tumor suppressor in UCs. The glycosylated-GREM1 hampered cell proliferation, migration, invasion, and in vitro angiogenesis through interaction with TGFB1 to inactivate TGFß/BMP/SMAD-mediated EMT in an autocrine manner.

Involvement of phorbol-12-myristate-13-acetate-induced protein 1 in goniothalamin-induced TP53-dependent and -independent apoptosis in hepatocellular carcinoma-derived cells.

The objective was to investigate the upstream apoptotic mechanisms that were triggered by a styrylpyrone derivative, goniothalamin (GTN), in tumor protein p53 (TP53)-positive and -negative hepatocellular carcinoma (HCC)-derived cells. Effects of GTN were evaluated by the flow cytometry, alkaline comet assay, immunocytochemistry, small-hairpin RNA interference, mitochondria/cytosol fractionation, quantitative reverse transcription-polymerase chain reaction, immunoblotting analysis and caspase 3 activity assays in two HCC-derived cell lines. Results indicated that GTN triggered phorbol-12-myristate-13-acetate-induced protein 1 (PMAIP1, also known as NOXA)-mediated apoptosis via TP53-dependent and -independent pathways. In TP53-positive SK-Hep1 cells, GTN furthermore induced TP53 transcription-dependent and -independent apoptosis. After GTN treatment, accumulation of reactive oxygen species, formation of DNA double-strand breaks, transactivation of TP53 and/or PMAIP1 gene, translocation of TP53 and/or PMAIP1 proteins to mitochondria, release of cytochrome c from mitochondria, cleavage of caspases and induction of apoptosis in both cell lines were sustained. GTN might represent a novel class of anticancer drug that induces apoptosis in HCC-derived cells through PMAIP1 transactivation regardless of the status of TP53 gene.

Vertically integrated diffractive gratings on photonic crystal surface emitting lasers.

We designed and fabricated a photonic crystal surface emitting laser (PCSEL) with vertically integrated diffractive optical elements on their top to study the mechanism of static beam steering on a single chip. The deflected output beam by the self-formed periodic ITO cladding layer of the PCSEL can be further steered by changing the grating period and azimuthal angle of the diffractive gratings relative to the photonic crystal. Through the analysis of photonic band structure and lasing characteristics, the periodic ITO structure is coupled to the photonic crystal band, whereas the integrated grating serves the diffractive function only. The findings pave the way for the design of PCSELs enabling single or multiple output beam with varying direction capability. This type of laser is regarded as an ideal light source for various applications, such as light detection and ranging and three-dimensional sensing systems.

Sustained-Release and pH-Adjusted Alginate Microspheres-Encapsulated Doxorubicin Inhibit the Viabilities in Hepatocellular Carcinoma-Derived Cells.

The objective of this study aimed to develop biodegradable calcium alginate microspheres carrying doxorubicin (Dox) at the micrometer-scale for sustained release and the capacity of pH regulatory for transarterial chemoembolization. Ultrasonic atomization and CaCl2 cross-linking technologies were used to prepare the microspheres. A 4-by-5 experiment was first designed to identify imperative parameters. The concentration of CaCl2 and the flow rate of the pump were found to be critical to generate microspheres with a constant volume median diameter (~39 µm) across five groups with different alginate: NaHCO3 ratios using each corresponding flow rate. In each group, the encapsulation efficiency was positively correlated to the Dox-loading %. Fourier-transform infrared spectroscopy showed that NaHCO3 and Dox were step-by-step incorporated into the calcium alginate microspheres successfully. Microspheres containing alginate: NaHCO3 = 1 exhibited rough and porous surfaces, high Young's modulus, and hardness. In each group with the same alginate: NaHCO3 ratio, the swelling rates of microspheres were higher in PBS containing 10% FBS compared to those in PBS alone. Microspheres with relatively high NaHCO3 concentrations in PBS containing 10% FBS maintained better physiological pH and higher accumulated Dox release ratios. In two distinct hepatocellular carcinoma-derived cell lines, treatments with microspheres carrying Dox demonstrated that the cell viabilities decreased in groups with relatively high NaHCO3 ratios in time- and dose-dependent manners. Our results suggested that biodegradable alginate microspheres containing relatively high NaHCO3 concentrations improved the cytotoxicity effects in vitro.

A protocol for differential staining of cartilages and ossified bones in fetal and adult mouse skeletons using alcian blue and alizarin red S.

The technique for clearing and staining whole specimens consists of many steps. This study discusses the alcian blue/alizarin red S staining method and aims to provide a useful reference and review for users who intend to do this staining. To specifically address the influences of tissue removal on staining results, the mouse fetuses at embryonic stage E18.5 and adult mice at 12 weeks of age were used in this study. The fetuses were divided into three groups: Group 1 skin, muscle, and viscera removed, Group 2 skin and muscle removed, and Group 3 viscera removed. For successful skeletal staining, it was concluded that (1) skin removal from fetuses was necessary for alcian blue staining but unnecessary for alizarin red S staining, (2) removal of muscle surrounding thorax and neck of fetuses could improve transparency effects, (3) retaining fetal viscera would not significantly affect transparency but might avoid the tissue damage, and (4) complete skin, muscle, and viscera removal were essential for good staining of adult mice. The representative images and detailed staining procedures might be good for researchers presently using alcian blue and alizarin red S staining to differentiate cartilages and ossified bones.

Effect of Passivation Layer on the Thin Film Perovskite Random Lasers.

Novel functionalities of disorder-induced scattering effect in random lasers, attributed to low spatial coherence, draw remarkable attention in high-contrast to superior quality speckle-free imaging applications. This paper demonstrates perovskite-polystyrene (PS)-based random lasing action with robust optical performance at room temperature. Optical characterizations are carried out upon perovskite thin films addition with polystyrene of different mixing concentrations (wt.%). A low threshold lasing operation is achieved with an increasing concentration of polystyrene, accompanying a wavy surface texture with high surface roughness. The rough surface dominating multiple scattering effects leads to enhanced feedback efficiency. Moreover, this study also elucidates efficient fabrication process steps for the development of high quality and durable PS-based random lasers. With the advantages of reduced coherent artifacts and low spatial coherence, speckle free projection images of the USAF (U. S. Air Force MIL-STD-150A standard of 1951) resolution test chart are shown for different PS-based random lasers.