Potato: from functional genomics to genetic improvement.

Potato is the most widely grown non-grain crop and ranks as the third most significant global food crop following rice and wheat. Despite its long history of cultivation over vast areas, slow breeding progress and environmental stress have led to a scarcity of high-yielding potato varieties. Enhancing the quality and yield of potato tubers remains the ultimate objective of potato breeding. However, conventional breeding has faced challenges due to tetrasomic inheritance, high genomic heterozygosity, and inbreeding depression. Recent advancements in molecular biology and functional genomic studies of potato have provided valuable insights into the regulatory network of physiological processes and facilitated trait improvement. In this review, we present a summary of identified factors and genes governing potato growth and development, along with progress in potato genomics and the adoption of new breeding technologies for improvement. Additionally, we explore the opportunities and challenges in potato improvement, offering insights into future avenues for potato research.

Glycyrrhizic acid attenuates the malignant biological properties of nonalcoholic fatty liver disease-related hepatocellular carcinoma.

Glycyrrhizic acid (GA) has effects on anti-hepatic fibrosis, anti-tumor and prevention from hepatocellular carcinoma (HCC) progression. Yet, the capacity of GA to ameliorate the advance of HCC pertinent to nonalcoholic fatty liver disease (NAFLD) remains to be clarified. We used the CCK-8 method to detect the optimal treatment concentration and time for L-02 cells, palmitic acid (PA)-induced L-02 cells and HepG2 cells, and selected 40 µM and 48 h to treat PA-induced L-02 cells and 60 µM for 24 h to treat HepG2 cells. Moreover, functional associations of HepG2 cells were elucidated through various assays. The results showed that GA demonstrated enhances lipid deposition and alleviates the inflammatory response in L-02 cells induced by palmitic acid. Simultaneously, we found that GA inhibits the proliferation, migration, and invasion while promoting apoptosis in HepG2 cells. In pursuit of constructing of HCC model rats, a combination of high-fat diets and diethylnitrosamine was utilized. The results showed that GA significantly decreased the liver index, body weight, liver weight, and the number of nodules in HCC model rats. Moreover, GA mitigated infiltration and heightened apoptosis in these rats. Mechanistically, GA notably attenuated the KKß/NF-κB pathway in both HepG2 cells and the HCC model rats. In conclusion, GA functions as an inhibitor in the progression of NAFLD-related HCC cells, which might be relevant to the KKß/NF-κB pathway. Therefore, GA is a potential drug for NAFLD-related HCC treatment.

WDR12/RAC1 axis promoted proliferation and anti-apoptosis in colorectal cancer cells.

BACKGROUND: WD repeat domain 12 (WDR12) plays a crucial role in the ribosome biogenesis pathway. However, its biological function in colorectal cancer (CRC) remains poorly understood. Therefore, this study aims to investigate the roles of WDR12 in the occurrence and progression of CRC, as well as its underlying mechanisms. METHODS: The expression of WDR12 was assessed through The Cancer Genome Atlas (TCGA) and the Human Protein Atlas (HPA) database. Functional experiments including Celigo assay, MTT assay, and Caspase-3/7 assay were conducted to validate the role of WDR12 in the malignant progression of CRC. Additionally, mRNA chip-sequencing and ingenuity pathway analysis (IPA) were performed to identify the molecular mechanism. RESULTS: WDR12 expression was significantly upregulated in CRC tissues compared to normal colorectal tissues. Knockdown of WDR12 reduced proliferation and promoted apoptosis of CRC cell lines in vitro and in vivo experiments. Furthermore, WDR12 expression had a significantly inverse association with diseases and functions, including cancer, cell cycle, DNA replication, recombination, cellular growth, proliferation and repair, as revealed by IPA analysis of mRNA chip-sequencing data. Moreover, the activation of cell cycle checkpoint kinases proteins in the cell cycle checkpoint control signaling pathway was enriched in the WDR12 knockdown CRC cell lines. Additionally, downregulation of rac family small GTPase 1 (RAC1) occurred upon WDR12 knockdown, thereby facilitating the proliferation and anti-apoptosis of CRC cells. CONCLUSION: Our study demonstrates that the WDR12/RAC1 axis promotes tumor progression in CRC. Therefore, WDR12 may serve as a novel oncogene and a potential target for individualized therapy in CRC. These findings provide an experimental foundation for the clinical development of drugs targeting the WDR12/RAC1 axis.

Intravoxel incoherent motion imaging used to assess tumor microvascular changes after transarterial chemoembolization in a rabbit VX2 liver tumor model.

Purpose: To evaluate the correlation between microvascular density (MVD) and intravoxel incoherent motion (IVIM) magnetic resonance imaging (MRI) parameters and the effect of glycolytic flux after transarterial chemoembolization (TACE) in a rabbit VX2 liver tumor. Materials and methods: VX2 liver tumor allografts in 15 New Zealand white rabbits were treated with sterile saline (control group, n = 5) or lipiodol-doxorubicin emulsion (experimental group, n = 10). MRI was performed 2 weeks after the procedure to evaluate IVIM parameters, including apparent diffusion coefficient (ADC), pure diffusion coefficient (D), pseudodiffusion coefficient (D*), and perfusion fraction (PF). All animal samples were taken of the tumor and surrounding liver. Immunostaining for CD31, CD34, CD105, and VEGF was used to evaluate MVD. The protein expression of Glut4, HK2, PKM2, LDHA, and MCT1 was determined using western blotting. Pearson correlation tests were used to analyze the relationship between MVD and IVIM parameters. Results: D* value in the peritumoral region was negatively correlated with CD34 (r = -0.71, P = 0.01). PF value positively correlated with CD34 (r = 0.68, P = 0.015), CD105 (r = 0.76, P = 0.004) and VEGF (r = 0.72, P = 0.008) in the peritumoral region. Glut4, HK2, PKM2, and MCT1 in the peritumoral regions were higher in the experimental group than in the control group (all P < 0.05). Conclusion: IVIM parameters were correlated with MVD in the intratumoral and peritumoral regions after TACE in a rabbit liver tumor model. The angiogenesis reflected by MVD may be related to changes of glycolytic flux.

Associations of muscle and adipose tissue parameters with long-term outcomes in middle and low rectal cancer: a retrospective cohort study.

OBJECTIVE: To investigate the role of preoperative body composition analysis for muscle and adipose tissue distribution on long-term oncological outcomes in patients with middle and low rectal cancer (RC) who received curative intent surgery. METHODS: A total of 155 patients with middle and low rectal cancer who underwent curative intent surgery between January 2014 and December 2016 were included for the final analysis. Skeletal muscle area (SMA), skeletal muscle radiodensity (SMD), visceral fat area (VFA) and mesorectal fat area (MFA) were retrospectively measured using preoperative CT images. To standardize the area according to patient stature, SMA was divided by the square of the height (m2) and the skeletal muscle mass index (SMI, cm2/m2) was obtained. Each median values of the distribution in male and female served as cut-off point for SMI, SMD, VFA, and MFA, respectively. Univariate and multivariate analysis were performed to evaluate the association between body composition and long-term oncological outcomes. Overall survival (OS) measured in months from the day of primary surgery until death for any cause. Disease-free survival (DFS) was defined as the interval between surgery and tumor recurrence. The Kaplan-Meier method with log-rank testing was used to validate prognostic biomarkers. Intraclass correlation coefficient (ICC) was used to evaluate interobserver and intraobserver reproducibility for SMA, SMD, MFA,VFA. RESULTS: During the follow-up period, 42 (27.1%) patients had tumor recurrence; 21 (13.5%) patients died. The sex-specific median value of SMI was 28.6 cm2/m2 for females and 48.2 cm2/m2 for males. The sex-specific median value of SMD was 34.7 HU for females and 37.4 HU for males. The sex-specific median value of VFA was 123.1 cm2 for females and 123.2 cm2 for males. The sex-specific median value of MFA was 13.8 cm2 for females and 16.0 cm2 for males. In the Cox regression multivariate analysis, SMI (P = 0.036), SMD (P = 0.022), and postoperative complications grades (P = 0.042) were significantly different between death group and non-death group; SMD (P = 0.011) and MFA (P = 0.022) were significantly different between recurrence group and non-recurrence group. VFA did not show any significant differences. By the Kaplan-Meier method with log-rank testing, DFS was significantly longer in patients with high-MFA (P = 0.028) and shorter in patients with low-SMD (P = 0.010), OS was significantly shorter in patients with low-SMI (P = 0.034) and low-SMD (P = 0.029). CONCLUSIONS: Quantitative evaluation of skeletal muscle mass and adipose tissue distributions at initial diagnosis were important predictors for long-term oncologic outcomes in RC patients. SMD and SMI were independent factors for predicting OS in patients with middle and low rectal cancer who had radical surgery. SMD and MFA were independent factors for predicting DFS in patients with middle and low rectal cancer who had radical surgery.

Activated Hepatic Stellate Cells Promote the M1 to M2 Macrophage Transformation and Liver Fibrosis by Elevating the Histone Acetylation Level.

Liver fibrosis results from the formation of fibrous scars of hepatic stellate cells by various chronic liver diseases. Considering that the liver is the most important metabolic organ in the human body, exploring the metabolic characteristics of liver fibrosis is expected to discover new markers and therapeutic targets. In this study, we first used mouse model to verify that both lactate content and histone acetylation levels were significantly increased in hepatic fibrosis mice. At the same time, it was confirmed that activated hepatic stellate cells (HSCs) cocultured with M1 macrophages can promote their transformation into M2 macrophages in hepatic stellate cell line and primary hepatic stellate cells. In addition, the addition of lactic acid to the medium in which M1 cells are cultured can promote their transformation into M2 macrophages. Therefore, we concluded that activated HSCs can promote the transformation of M1 to M2 macrophages through lactate accumulation, thereby causing liver fibrosis.

Regulatory Mechanism miR-302a-3p/E2F1/SNHG3 Axis in Nerve Repair Post Cerebral Ischemic Stroke.

OBJECTIVE: Cerebral ischemic stroke (CIS) remains a primary cause of death worldwide. The current knowledge has identified the implication of microRNAs (miRNAs) in the pathophysiology of CIS. This study investigated the mechanism of miR-302a-3p in nerve repair post CIS. METHODS: A middle cerebral artery occlusion (MCAO) model was established in mice to simulate CIS. miR-302a-3p expression in brain tissues of MCAO mice was up-regulated by injecting agomiR-302a-3p. Neurological deficits of MCAO mice were evaluated through neurological function score, forelimb placing test, and balance beam walking test. Neuronal damage was measured using Nissl staining. The concentrations of nerve injury-related factors (S100B and GFAP) and the contents of neuroinflammatory factors (TNF-α and IL-1ß) in serum were examined using ELISA kits. miR-302a-3p, E2F1, and long non-coding RNA (lncRNA) SNHG3 expressions in brain tissues of MCAO mice were determined using RT-qPCR and Western blot. The binding relationships between miR-302a-3p and E2F1 and E2F1 and SNHG3 were validated using dual-luciferase and ChIP assays, respectively. RESULTS: miR-302a-3p expression was reduced in brain tissues of MCAO mice. miR-302a-3p overexpression increased the number of neurons, decreased the concentrations of S100B and GFAP, reduced the contents of TNF-α and IL-1ß, promoted nerve repair, and alleviated CIS-induced brain injury. miR-302a-3p targeted E2F1 expression, and E2F1 activated SNHG3 transcription. E2F1 overexpression or SNHG3 overexpression reversed the effect of miR-302a-3p overexpression on nerve repair in MCAO mice. CONCLUSION: miR-302a-3p overexpression repressed SNHG3 transcription by targeting E2F1 expression, thereby promoting nerve repair and alleviating CIS.

Dual Dynamic Scheduling for Hierarchical QoS in Uplink-NOMA: A Reinforcement Learning Approach.

The demand for bandwidth-intensive and delay-sensitive services is surging daily with the development of 5G technology, resulting in fierce competition for scarce radio resources. Power domain Nonorthogonal Multiple Access (NOMA) technologies can dramatically improve system capacity and spectrum efficiency. Unlike existing NOMA scheduling that mainly focuses on fairness, this paper proposes a power control solution for uplink hybrid OMA and PD-NOMA in dual dynamic environments: dynamic and imperfect channel information together with the random user-specific hierarchical quality of service (QoS). This paper models the power control problem as a nonconvex stochastic, which aims to maximize system energy efficiency while guaranteeing hierarchical user QoS requirements. Then, the problem is formulated as a partially observable Markov decision process (POMDP). Owing to the difficulty of modeling time-varying scenes, the urgency of fast convergency, the adaptability in a dynamic environment, and the continuity of the variables, a Deep Reinforcement Learning (DRL)-based method is proposed. This paper also transforms the hierarchical QoS constraint under the NOMA serial interference cancellation (SIC) scene to fit DRL. The simulation results verify the effectiveness and robustness of the proposed algorithm under a dual uncertain environment. As compared with the baseline Particle Swarm Optimization algorithm (PSO), the proposed DRL-based method has demonstrated satisfying performance.

The protective role of the miR-25-mediated notch signaling pathway in the memory capacity and brain tissue of mice with central nervous system infections.

OBJECTIVE: The study aimed to explore the role of miR-25 and the notch signaling pathway in the memory capacity and brain tissue of mice with central nervous system (CNS) infections. METHODS: A bioinformatics website and the dual-luciferase reporter assay were used to analyze the targeting relationship between miR-25 and Notch1. The mice were randomized into 7 groups (n=10 per group), including the normal group, the model group (lipopolysaccharide at a dose of 500 µg/kg for the model establishment), the NC group, the miR-25 mimic group, the miR-25 inhibitor group, the DAPT group, and the miR-25 inhibitor + DAPT group. qRT-PCR and western blot were used to measure the miR-25, Notch1, and Hes5 expression levels in the hippocampal CA1 region of the mice's brains, along with the cyclo-oxygenase 2 (COX-2) and inducible nitric oxide synthase (iNOS) levels in the mice's hippocampi. RESULTS: Compared with the normal mice, the model mice had up-regulated miR-25, COX-2, and iNOS expressions and down-regulated Notch1 and Hes5 expressions, lower superoxide dismutase (SOD) levels in the hippocampi, and higher malondialdehyde (MDA) levels. Compared with the model group, the miR-25 mimic and DAPT groups had down-regulated Notch1 and Hes5 expressions, lower learning and memory capacities and SOD levels, higher MDA levels, and up-regulated COX-2 and iNOS expressions. CONCLUSION: Down-regulating miR-25 may improve the memory capacity in mice with CNS infections by activating the Notch signaling pathway.

Clinical and X-ray characteristics for expressions of different receptors in patients with breast cancer. / ä¸åŒå—ä½“è¡¨è¾¾çš„ä¹³è ºç™Œæ‚£è€ çš„ä¸´åºŠä¸ŽX线特征.

OBJECTIVES: Clarifying the expression of breast cancer receptor is the key to clinical treatment for breast cancer. This study aims to explore the correlation between X-ray and clinical characteristics of 4 molecular subtypes and their receptor types of breast cancer. METHODS: A total of 439 breast cancer patients who confirmed by pathology and performed X-ray examination were enrolled. The X-ray and clinical characteristics of 4 molecular subtypes and the expression of their receptors were analyzed. RESULTS: Luminal A type showed the highest proportion of spiculate masses, and the lowest calcification score, showing significant difference with other 3 subtypes (all P<0.001). The age in the human epidermal growth factor 2 (HER2) overexpression type group was older, the proportions of menopause, the calcification score, and the calcification score with 9-12 were higher, the sizes of the tumor were greater in the HER2 overexpression type group than those in the other 3 molecular subtype groups (age P<0.05, the rest P<0.01). The proportions of regular shape, edge indistinct, and high-grade invasive ductal carcinoma in the triple-negative type group were higher than those in the other 3 molecular subtype groups (all P<0.001). The proportions of non-menopausal patients and spiculate tumors in the estrogen receptor (ER) positive and/or progesterone receptor (PR) positive groups were higher than those in both ER and PR negative group (P<0.001 and P=0.001, respectively). The proportions of calcification fraction and high-grade invasive ductal carcinoma were higher, tumor sizes were greater in the HER2 positive group, Ki-67≥20% group than those in the HER2 negative group, Ki-67<20% group, respectively (P<0.001 or P<0.05, respectively). CONCLUSIONS: Four molecular subtypes of breast cancer and their receptor expressions are correlated with X-ray and clinical characteristics, which can provide a basis for clinical diagnosis and treatment.

Mapping quantitative trait loci and predicting candidate genes for leaf angle in maize.

Leaf angle of maize is a fundamental determinant of plant architecture and an important trait influencing photosynthetic efficiency and crop yields. To broaden our understanding of the genetic mechanisms of leaf angle formation, we constructed a F3:4 recombinant inbred lines (RIL) population to map QTL for leaf angle. The RIL was derived from a cross between a model inbred line (B73) with expanded leaf architecture and an elite inbred line (Zheng58) with compact leaf architecture. A sum of eight QTL were detected on chromosome 1, 2, 3, 4 and 8. Single QTL explained 4.3 to 14.2% of the leaf angle variance. Additionally, some important QTL were confirmed through a heterogeneous inbred family (HIF) approach. Furthermore, twenty-four candidate genes for leaf angle were predicted through whole-genome re-sequencing and expression analysis in qLA02-01and qLA08-01 regions. These results will be helpful to elucidate the genetic mechanism of leaf angle formation in maize and benefit to clone the favorable allele for leaf angle. Besides, this will be helpful to develop the novel maize varieties with ideal plant architecture through marker-assisted selection.

Involvement of prenucleation clusters in calcium phosphate mineralization of collagen.

Involvement of thermodynamically-stable prenucleation clusters (PNCs) in the biomineralization of collagen has been speculated since their existence was reported in mineralization systems. It has been hypothesized that intrafibrillar mineralization proceeds via nucleation of inhibitor-stabilized intermediates produced by liquid-liquid separation (aka. polymer-induced liquid precursors; PILPs). Here, the contribution of PNCs and PILPs to calcium phosphate intrafibrillar mineralization of collagen was examined in a model with a semipermeable membrane that excludes nucleation inhibitor-stabilized PILPs from reaching the collagen fibrils, using cryogenic electron microscopy of reconstituted fibrils and conventional transmission electron microscopy of collagen sponges. Molecular dynamics simulation with the Interface force field (IFF) was used to confirm the existence of PILPs with amorphous calcium phosphate and elucidate details of the dynamics. Furthermore, intrafibrillar mineralization of single collagen fibrils was experimentally observed with unstabilized PNCs when anionic/cationic polyelectrolytes were used to establish Donnan equilibrium across the semipermeable membrane. Molecular dynamics simulation verified PNC formation within the collagen intrafibrillar gap zones at the atomic scale and explained the role of external PILPs. The PILPs decrease the interfibrillar water content and increase the interfibrillar ionic concentration. Nevertheless, intrafibrillar mineralization of collagen sponges with PNCs alone was inefficacious, being constrained by competition from extrafibrillar mineral precipitation. STATEMENT OF SIGNIFICANCE: Compared with conventional PILP-based intrafibrillar mineralization, mineralization of collagen fibrils using unstabilized PNCs is constrained by competition from extrafibrillar mineral deposition. The narrow window of opportunity for PNCs to produce intrafibrillar mineralization provides a plausible explanation for the feasibility of nucleation inhibitor-free intrafibrillar apatite assembly during reconstitution of type I collagen.

15-hydroxy-6α,12-epoxy-7ß,10αH,11ßH-spiroax-4-ene-12-one exerts anti-tumor effects against osteosarcoma through apoptosis induction.

Osteosarcoma is the most common type of malignant bone tumor, which has an overall survival rate of only 15-30%. The present study aimed to investigate the effects of 15-hydroxy-6α,12-epoxy-7ß,10αH,11ßH-spiroax-4-ene-12-one (HESEO), a compound extracted from the endophytic fungus Penicillium sp. FJ-1 isolated from Avicennia marina, on the proliferation of osteosarcoma cells and to explore its underlying mechanisms of action. Cell number was counted to measure the cell proliferation. JC-1 reagent was used to measure mitochondrial membrane potential. ELISA was used to measure the cytochrome c level and caspase activities. Apoptosis was detected by Annexin V-Propidium Iodide staining. Gene and protein expression were measured by reverse-transcription-PCR and western blot analysis, respectively. Additionally, the anti-tumor effects of HESEO were explored within a syngeneic osteosarcoma tumor model. The results suggested that HESEO significantly inhibited the proliferation of osteosarcoma cells and induced apoptosis of MG-63 cells, evidenced by their decreased mitochondrial membrane potential, and increased cytochrome c release, caspase activities and percentage of apoptotic cells. In addition, HESEO increased the expression of pro-apoptotic genes and proteins compared with control cells. The results indicated that HESEO may act through increasing p53 upregulated modulator of apoptosis expression. Furthermore, HESEO treatment significantly increased the survival time and decreased the tumor burden of osteosarcoma tumor-bearing mice compared with vehicle treatment. Furthermore, combined treatment with HESEO enhanced the effects of the chemotherapeutic agent methotrexate on a lung metastasis osteosarcoma model. These data suggested that HESEO could be developed as a potential anti-tumor agent against osteosarcoma.

Bone mineral density measurement combined with vertebral fracture assessment increases diagnosis of osteoporosis in postmenopausal women.

OBJECTIVE: Osteoporosis is diagnosed based on the results of BMD assessment and/or fragility fractures. Vertebral fracture is the most common fragility fracture. Many vertebral fractures are asymptomatic and are not clinically recognized. Early detection of vertebral fracture may increase diagnosis of osteoporosis. In this study, we performed BMD measurement combined with vertebral fracture assessment (VFA) by DXA for the postmenopausal women receiving the first bone densitometry and studied the impact of VFA on the diagnosis of osteoporosis. METHODS: A total of 502 postmenopausal women were enrolled in our study. Patients' age was 66.7 ± 9.5 years. All patients had BMD assessment and VFA by dual-energy X-ray absorptiometry. Genant's semiquantitative assessment was used. The impact of VFA on the diagnosis of osteoporosis was studied. All parameters of groups were compared using the Chi-squared test. RESULTS: There were 257 patients with T-score ≤-2.5, 202 patients with a T-score between -1 and - 2.5, and 43 patients with BMD within the normal range. There were 162 patients with 345 fractured vertebrae identified by VFA, among which 84% of patients were previously unknown. Osteoporosis or severe osteoporosis was presented in 51.2% patients diagnosed by BMD alone, in 55.2% patients diagnosed by BMD plus fracture history, and in 62.4% of patients diagnosed by BMD plus fracture history and VFA. Severe osteoporosis significantly increased by 17.2% in patients receiving VFA. CONCLUSIONS: VFA combined with BMD can detect previously unknown vertebral fractures and increase clinical diagnosis of osteoporosis. It is plausible to speculate that this method should be considered in postmenopausal women for the first BMD assessment.

Hollow mesoporous zirconia delivery system for biomineralization precursors.

Strategies based on the combination of nanocarrier delivery systems and scaffolds provide bone tissue engineering scaffolds with multifunctional capability. Zirconia, a biocompatible ceramic commonly used in orthopedic and dental implants, was used to synthesize hollow mesoporous nanocapsules for loading, storage and sustained release of a novel polyamine-stabilized liquid precursor phase of amorphous calcium phosphate (PAH-ACP) for collagen biomineralization and bone marrow stromal cells osteoinduction. Hollow mesoporous zirconia (hmZrO2) nanocapsules loaded with biomimetic precursors exhibited pH-sensitive release capability and good biocompatibility. The PAH-ACP released from loaded hmZrO2 still retained the ability to infiltrate and mineralize collagen fibrils as well as exhibited osteoinductivity. A collagen scaffold blended with PAH-ACP@hmZrO2 supplement and stem cells may be a promising tool for bone tissue engineering. STATEMENT OF SIGNIFICANCE: The advent of nanotechnology has catalyzed the development of bone tissue engineering strategies based on the combination of nanocarrier delivery systems and scaffolds, which provide distinct advantages, including the possibilities of sustained release and protection of the bioactive agents, site-specific pharmacological effects and reduction of side effects. Herein, hollow mesoporous zirconia (hmZrO2) nanocapsules with pH-sensitive capacity were synthesized for loading, storage and sustained release of a novel polyamine-stabilized liquid precursor phase of ACP (PAH-ACP). The loaded nanocapsules show good biocompatibility and demonstrate bioactivities for collagen biomineralization and bone marrow stromal cells osteoinduction. Our results may offer a promising tool for designing bone tissue engineering "cocktail therapy" involving seeding scaffolds with biomineralization precursors loaded hmZrO2 supplement and stem cells.

Mechanism of bioactive molecular extraction from mineralized dentin by calcium hydroxide and tricalcium silicate cement.

OBJECTIVES: The objective of the present study was to elucidate the mechanism of bioactive molecule extraction from mineralized dentin by calcium hydroxide (Ca(OH)2) and tricalcium silicate cements (TSC). METHODS AND RESULTS: Transmission electron microscopy was used to provide evidence for collagen degradation in dentin surfaces covered with Ca(OH)2 or a set, hydrated TSC for 1-3 months. A one micron thick collagen degradation zone was observed on the dentin surface. Fourier transform-infrared spectroscopy was used to identify increases in apatite/collagen ratio in dentin exposed to Ca(OH)2. Using three-point bending, dentin exposed to Ca(OH)2 exhibited significant reduction in flexural strength. Using size exclusion chromatography, it was found that the small size of the hydroxyl ions derived from Ca(OH)2 enabled those ions to infiltrate the intrafibrillar compartment of mineralized collagen and degrade the collagen fibrils without affecting the apatite minerals. Using ELISA, TGF-ß1 was found to be extracted from dentin covered with Ca(OH)2 for 3 months. Unlike acids that dissolve the mineral component of dentin to release bioactive molecules, alkaline materials such as Ca(OH)2 or TSC released growth factors such as TGF-ß1 via collagen degradation. SIGNIFICANCE: The bioactive molecule extraction capacities of Ca(OH)2 and TSC render these dental materials excellent for pulp capping and endodontic regeneration. These highly desirable properties, however, appear to be intertwined with the untoward effect of degradation of the collagen matrix within mineralized dentin, resulting in reduced flexural strength.

Primum non nocere - The effects of sodium hypochlorite on dentin as used in endodontics.

The medical literature is replete with the maxim 'primum non nocere', cautioning health care providers to avoid doing any harm to human subjects in their delivery of medical care. Sodium hypochlorite (NaOCl) is a well-established irrigant for root canal treatment because of its antimicrobial and organic tissue remnant dissolution capability. However, little is known about the deleterious effect of this strong oxidizing agent on the integrity of human mineralized dentin. Iatrogenically-induced loss of dentin integrity may precipitate post-treatment root fracture and has potential medico-legal complications. In the present work, transmission electron microscopy provided evidence for collagen destruction in the surface/subsurface of dentin treated with high NaOCl concentrations and long contact times. Size exclusion chromatography showed that the hypochlorite anion, because of its small size, penetrated the water compartments of apatite-encapsulated collagen fibrils, degraded the collagen molecules and produced a 25-35µm thick, non-uniform "ghost mineral layer" with enlarged, coalesced dentinal tubules and their lateral branches. Fourier transform-infrared spectroscopy identified increases in apatite/collagen ratio in NaOCl-treated dentin. The apatite-rich, collagen-sparse dentin matrix that remained after NaOCl treatment is more brittle, as shown by the reductions in flexural strength. Understanding the deleterious effects of NaOCl on mineralized dentin enables one to balance the risks and benefits in using high NaOCl concentrations for lengthy periods in root canal debridement. Delineating the mechanism responsible for such a phenomenon enables high molecular weight, polymeric antimicrobial and tissue dissolution irrigants to be designed that abides by the maxim of 'primum non nocere' in contemporary medical practices. STATEMENT OF SIGNIFICANCE: The antimicrobial and tissue-dissolution capacities of NaOCl render it a well-accepted agent for root canal debridement. These highly desirable properties, however, appear to be intertwined with the untoward effect of collagen matrix degradation within mineralized dentin. Because of its small size, the hypochlorite anion is capable of infiltrating mineralized collagen and destroying the collagen fibrils, producing a mineral-rich, collagen sparse ghost mineral matrix with reduced flexural strength. Findings from the present work challenge the biosafety of NaOCl when it is used in high concentrations and for lengthy time periods during root canal treatment, and laid the background work for future biomaterials design in debridement of the canal space.

No-waiting dentine self-etch concept-Merit or hype.

OBJECTIVE: A recently-launched universal adhesive, G-Premio Bond, provides clinicians with the alternative to use the self-etch technique for bonding to dentine without waiting for the adhesive to interact with the bonding substrate (no-waiting self-etch; Japanese brochure), or after leaving the adhesive undisturbed for 10s (10-s self-etch; international brochure). The present study was performed to examine in vitro performance of this new universal adhesive bonded to human coronal dentine using the two alternative self-etch modes. METHODS: One hundred and ten specimens were bonded using two self-etch application modes and examined with or without thermomechanical cycling (10,000 thermal cycles and 240,000 mechanical cycles) to simulate one year of intraoral functioning. The bonded specimens were sectioned for microtensile bond testing, ultrastructural and nanoleakage examination using transmission electron microscopy. Changes in the composition of mineralised dentine after adhesive application were examined using Fourier transform infrared spectroscopy. RESULTS: Both reduced application time and thermomechanical cycling resulted in significantly lower bond strengths, thinner hybrid layers, and significantly more extensive nanoleakage after thermomechanical cycling. Using the conventional 10-s application time improved bonding performance when compared with the no-waiting self-etch technique. Nevertheless, nanoleakage was generally extensive under all testing parameters employed for examining the adhesive. CONCLUSION: Although sufficient bond strength to dentine may be achieved using the present universal adhesive in the no-waiting self-etch mode that does not require clinicians to wait prior to polymerisation of the adhesive, this self-etch concept requires further technological refinement before it can be recommended as a clinical technique. CLINICAL SIGNIFICANCE: Although the surge for cutting application time to increase user friendliness remains the most frequently sought conduit for advancement of dentine bonding technology, the use of the present universal adhesive in the no-waiting self-etch mode may not represent the best use of the adhesive.

Biodegradable mesoporous delivery system for biomineralization precursors.

Scaffold supplements such as nanoparticles, components of the extracellular matrix, or growth factors have been incorporated in conventional scaffold materials to produce smart scaffolds for tissue engineering of damaged hard tissues. Due to increasing concerns on the clinical side effects of using large doses of recombinant bone-morphogenetic protein-2 in bone surgery, it is desirable to develop an alternative nanoscale scaffold supplement that is not only osteoinductive, but is also multifunctional in that it can perform other significant bone regenerative roles apart from stimulation of osteogenic differentiation. Because both amorphous calcium phosphate (ACP) and silica are osteoinductive, a biodegradable, nonfunctionalized, expanded-pore mesoporous silica nanoparticle carrier was developed for loading, storage, and sustained release of a novel, biosilicification-inspired, polyamine-stabilized liquid precursor phase of ACP for collagen biomineralization and for release of orthosilicic acid, both of which are conducive to bone growth. Positively charged poly(allylamine)-stabilized ACP (PAH-ACP) could be effectively loaded and released from nonfunctionalized expanded-pore mesoporous silica nanoparticles (pMSN). The PAH-ACP released from loaded pMSN still retained its ability to infiltrate and mineralize collagen fibrils. Complete degradation of pMSN occurred following unloading of their PAH-ACP cargo. Because PAH-ACP loaded pMSN possesses relatively low cytotoxicity to human bone marrow-derived mesenchymal stem cells, these nanoparticles may be blended with any osteoconductive scaffold with macro- and microporosities as a versatile scaffold supplement to enhance bone regeneration.

Mineralogenic characteristics of osteogenic lineage-committed human dental pulp stem cells following their exposure to a discoloration-free calcium aluminosilicate cement.

OBJECTIVES: An experimental discoloration-free calcium aluminosilicate cement has been developed with the intention of maximizing the beneficial attributes of tricalcium silicate cements and calcium aluminate cements. The present study examined the effects of this experimental cement (Quick-Set2) on the mineralogenic characteristics of osteogenic lineage-committed human dental pulp stem cells (hDPSCs), by comparing the cellular responses with a commercially available tricalcium silicate cement (white mineral trioxide aggregate (ProRoot(®) MTA); WMTA). METHODS: The osteogenic potential of hDPSCs exposed to the cements was examined using qRT-PCR for osteogenic gene expressions, Western blot for osteogenic-related protein expressions, alkaline phosphatase enzyme activity, Alizarin red S staining, Fourier transform infrared spectroscopy and transmission electron microscopy of extracellular calcium deposits. RESULTS: Results of the six assays indicated that osteogenic differentiation of hDPSCs was significantly enhanced after exposure to the tricalcium silicate cement or the experimental calcium aluminosilicate cement, with the former demonstrating better mineralogenic stimulation capacity. SIGNIFICANCE: The better osteogenic stimulating effect of the tricalcium silicate cement on hDPSCs may be due to its relatively higher silicate content, or higher OH(-) and Ca(2+) release. Further investigations with the use of in vivo animal models are required to validate the potential augmenting osteogenic effects of the experimental discoloration-free calcium aluminosilicate cement.