Temporal and Within-Sporophyte Variations in Triphenyltin Chloride (TPTCL) and Its Degradation Products in Cultivated Undaria pinnatifida.

Undaria pinnatifida can effectively deal with organotin pollution through its excellent accumulation and degradation capabilities found under laboratory conditions. However, nothing is known regarding its accumulation, degradation performance, and related impact factors in the wild farming area. In this study, we monitored triphenyltin chloride (TPTCL) contents and degradation products in different algal parts (blades, stipes, sporophylls, and holdfasts) of cultivated U. pinnatifida from December 2018 to May 2019. Our results showed that sporophytes had an accumulation and degradation capacity for TPTCL. The TPTCL contents and degradation products varied with the algal growth stages and algal parts. TPTCL accumulated in the blades at the growth stage and the blades, stipes, sporophylls, and holdfasts at the mature stage. The TPTCL content among algal parts was blades (74.92 ± 2.52 µg kg-1) > holdfasts (62.59 ± 1.42 µg kg-1) > sporophylls (47.24 ± 1.41 µg kg-1) > stipes (35.53 ± 0.55 µg kg-1). The primary degradation product DPTCL accumulated only in the blades at any stage, with a concentration of 69.30 ± 3.89 µg kg-1. The secondary degradation product MPTCL accumulated in the blades at the growth stage and in the blades, stipe, and sporophyll at the mature stage. The MPTCL content among algal parts was blades (52.80 ± 3.48 µg kg-1) > sporophylls (31.08 ± 1.53 µg kg-1) > stipes (20.44 ± 0.85 µg kg-1). The accumulation pattern of TPTCL and its degradation products seems closely related to nutrient allocation in U. pinnatifida. These results provide the basis for applying cultivated U. pinnatifida in the bioremediation of organotin pollution and the food safety evaluation of edible algae.

Screening of reference genes in tiger puffer (Takifugu rubripes) across tissues and under different nutritional conditions.

The present study was aimed at screening suitable reference genes for quantitative real-time polymerase chain reaction (qRT-PCR) in tiger puffer (Takifugu rubripes), an important aquaculture species in Asia and also a good model species for lipid research. Specifically, this reference gene screening was targeted at standardization of gene expression in different tissues (liver, muscle, brain, intestine, heart, eye, skin, and spleen) or under different nutritional conditions (starvation and different dietary lipid levels). Eight candidate reference genes (ribosomal protein L19 and L13 (RPL19 and RPL13), elongation factor-1 alpha (EF1α), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), hypoxanthine guanine phosphoribosyl transferase1 (HPRT1), beta-2-Microglobulin (B2M), 18S ribosomal RNA (18SrRNA), and beta actin (ACTB)) were evaluated with four algorithms (geNorm, NormFinder, BestKeeper, and comparative ΔCt method). The results showed that different algorithms generated inconsistent results. Based on these findings, RPL19, EF1α, 18SrRNA, and RPL13 were relatively stable in different tissues of tiger puffer. During starvation conditions, ACTB/RPL19 was the best reference gene combination. Under different dietary lipid levels, ACTB/RPL13 was the most suitable reference gene combination. The present results will help researchers to obtain more accurate results in future qRT-PCR analysis in tiger puffer.

First High-Density Linkage Map and QTL Fine Mapping for Growth-Related Traits of Spotted Sea bass (Lateolabrax maculatus).

Possessing powerful adaptive capacity and a pleasant taste, spotted sea bass (Lateolabrax maculatus) has a broad natural distribution and is one of the most popular mariculture fish in China. However, the genetic improvement program for this fish is still in its infancy. Growth is the most economically important trait and is controlled by quantitative trait loci (QTL); thus, the identification of QTLs and genetic markers for growth-related traits is an essential step for the establishment of marker-assisted selection (MAS) breeding programs. In this study, we report the first high-density linkage map of spotted sea bass constructed by sequencing 333 F1 generation individuals in a full-sib family using 2b-RAD technology. A total of 6883 SNP markers were anchored onto 24 linkage groups, spanning 2189.96 cM with an average marker interval of 0.33 cM. Twenty-four growth-related QTLs, including 13 QTLs for body weight and 11 QTLs for body length, were successfully detected, with phenotypic variance explained (PVE) ranging from 5.1 to 8.6%. Thirty potential candidate growth-related genes surrounding the associated SNPs were involved in cell adhesion, cell proliferation, cytoskeleton reorganization, calcium channels, and neuromodulation. Notably, the fgfr4 gene was detected in the most significant QTL; this gene plays a pivotal role in myogenesis and bone growth. The results of this study may facilitate marker-assisted selection for breeding populations and establish the foundation for further genomic and genetic studies investigating spotted sea bass.

Effects of elevated pCO2 and nutrient enrichment on the growth, photosynthesis, and biochemical compositions of the brown alga Saccharina japonica (Laminariaceae, Phaeophyta).

Ocean acidification and eutrophication are two major environmental issues affecting kelp mariculture. In this study, the growth, photosynthesis, and biochemical compositions of adult sporophytes of Saccharina japonica were evaluated at different levels of pCO2 (400 and 800 µatm) and nutrients (nutrient-enriched and non-enriched seawater). The relative growth rate (RGR), net photosynthetic rate, and all tested biochemical contents (including chlorophyll (Chl) a, Chl c, soluble carbohydrates, and soluble proteins) were significantly lower at 800 µatm than at 400 µatm pCO2. The RGR and the contents of Chl a and soluble proteins were significantly higher under nutrient-enriched conditions than under non-enriched conditions. Moreover, the negative effects of the elevated pCO2 level on the RGR, net photosynthetic rate, Chl c and the soluble carbohydrates and proteins contents were synergized by the elevated nutrient availability. These results implied that increased pCO2could suppress the growth and biochemical composition of adult sporophytes of S. japonica. The interactive effects of ocean acidification and eutrophication constitute a great threat to the cultivation of S. japonica due to growth inhibition and a reduction in quality.

The early repigmentation pattern of vitiligo is related to the source of melanocytes and by the choice of therapy: a retrospective cohort study.

BACKGROUND: Patients with vitiligo present with different repigmentation patterns in the early recovery stage. OBJECTIVES: To analyze the relationships between early repigmentation patterns in vitiliginous patches, their clinical characteristics, and therapeutic choices. METHODS: Patients with vitiligo seen in the Department of Dermatology, The First Affiliated Hospital of Nanjing Medical University from 2010 to 2015, were included, and their clinical records, especially photographs and medical treatments, were reviewed. RESULTS: One hundred and sixteen patients were included in this study, and 326 lesions with different degrees of depigmentation, locations, stages, distributions, therapies, and repigmentation patterns were included and analyzed. Perifollicular repigmentation occurred more frequently in lesions with complete depigmentation (P = 0.005), in non-sun exposed areas (P < 0.001), a stable stage (P = 0.008), and lesions treated with narrow band ultraviolet B (NB-UVB) (P < 0.001, despite lesion distributions). Marginal repigmentation is more frequent in lesions with complete depigmentation (P = 0.016), lesions treated without NB-UVB (P = 0.002), and facial lesions treated with topical vitamin D analogs (TVDAs) monotherapy (P = 0.022). Diffuse repigmentation is the predominant pattern in lesions with incomplete depigmentation (P < 0.001), in sun-exposed areas (P < 0.001), progressive stage (P = 0.044), and truncal lesions treated with TVDAs (P < 0.001). CONCLUSIONS: The different repigmentation patterns of vitiligo lesions depend on the different source and status of melanocytes and their abilities to produce melanin on the choice of therapy.

Protective effect of madecassoside on H2O2-induced oxidative stress and autophagy activation in human melanocytes.

BACKGROUND: Centella asiatica (L.) Urb. is a traditional Chinese medicine that has many medical applications, including wound healing and anti-oxidation. Some traditional Chinese Medicine doctors have found that it has therapeutic effects for external use in the repigmentation of vitiligo and post-inflammatory hypopigmentation. This study was designed to evaluate the effects of madecassoside, a major bioactive component of C. asiatica, on oxidative stress in human melanocytes and its possible mechanism of action. RESULTS: In H2O2-induced oxidative conditions, madecassoside inhibited melanocyte dendrite retraction, improved MMP and reduced the accumulation of [Ca2+]i in a concentration-dependent manner. Observations by TEM showed that madecassoside attenuated the damage of mitochondria in human melanocytes caused by oxidative stress. Furthermore, autophagy activation was demonstrated by AO staining and an increased LC3-II/LC3-I ratio. MATERIALS AND METHODS: Normal human melanocytes were treated with 0.01 mM H2O2 and varying concentrations of madecassoside (0, 10, 50, 100 µg/mL). Subsequently, the retraction velocity of melanocyte dendrites was assessed. Determination of mitochondrial membrane potential (MMP, ΔΨm) was performed by flow cytometry and intracellular calcium ([Ca2+]i) level were measured. Alterations of mitochondrial ultrastructure were observed by transmission electron microscopy (TEM). Acridine orange (AO) staining was used to measure autophagy. The LC3-II/LC3-I ratio, an indicator of autophagosome formation, was analyzed by western blot. CONCLUSIONS: These results demonstrate the antioxidative effect of madecassoside on human melanocytes subjected to oxidative damage via the activation of autophagy. Moreover, madecassoside could be a promising treatment for vitiligo mainly caused by oxidative stress.

Differences in the melanosome distribution within the epidermal melanin units and its association with the impairing background of leukoderma in vitiligo and halo nevi: a retrospective study.

Skin color is determined by the number of melanin granules produced by melanocytes that are transferred to keratinocytes. Melanin synthesis and the distribution of melanosomes to keratinocytes within the epidermal melanin unit (EMU) within the skin of vitiligo patients have been poorly studied. The ultrastructure and distribution of melanosomes in melanocytes and surrounding keratinocytes in perilesional vitiligo and normal skin were investigated using transmission electron microscopy (TEM). Furthermore, we performed a quantitative analysis of melanosome distribution within the EMUs with scatter plot. Melanosome count within keratinocytes increased significantly compared with melanocytes in perilesional stable vitiligo (P < 0.001), perilesional halo nevi (P < 0.01) and the controls (P < 0.01), but not in perilesional active vitiligo. Furthermore, melanosome counts within melanocytes and their surrounding keratinocytes in perilesional active vitiligo skin decreased significantly compared with the other groups. In addition, taking the means-standard error of melanosome count within melanocytes and keratinocytes in healthy controls as a normal lower limit, EMUs were graded into 3 stages (I-III). Perilesional active vitiligo presented a significantly different constitution in stages compared to other groups (P < 0.001). The distribution and constitution of melanosomes were normal in halo nevi. Impaired melanin synthesis and melanosome transfer are involved in the pathogenesis of vitiligo. Active vitiligo varies in stages and in stage II, EMUs are slightly impaired, but can be resuscitated, providing a golden opportunity with the potential to achieve desired repigmentation with an appropriate therapeutic choice. Adverse milieu may also contribute to the low melanosome count in keratinocytes.

Hashimoto's thyroiditis could be secondary to vitiligo: the possibility of antigen crossover and oxidative stress between the two diseases.

Autoimmune thyroid diseases (AITDs) are often accompanied by vitiligo, and the sera of patients with vitiligo often demonstrate increased frequencies of thyroid autoantibodies. In this study, we investigated the expression of melanocyte-associated antigens in tissues from patients with Hashimoto's thyroiditis (HT) without vitiligo using immunohistochemistry. Tissues of HT without vitiligo, as well as normal thyroid tissues, were both negative for the expression of NKI/beteb, gp100, tyrosinase-related protein 1 (TRP1), HMB-45 and S100, whereas they were positive for the expression of tyrosinase-related protein 2 (TRP2), lysosome-associated membrane protein 1 (LAMP1) and CD69. Tyrosinase (TYR) was only detected in tissues of HT, and levels of LAMP1 and CD69 were higher in tissues of HT than in normal thyroid tissues (p < 0.005). These results suggest the possibility of antigen crossover and oxidative stress between vitiligo and HT that might represent an immunological basis for secondary HT associated with vitiligo.

rhCSF3 accelerates the proliferation of human melanocytes in culture through binding CSF3R and the expression of CSF3R transcripts.

Melanogenic paracrine and autocrine cytokine networks have recently been discovered in vitro between melanocytes and other types of skin cells. Granulocyte colony-stimulating factor receptor (CSF3R) controls the survival, proliferation and differentiation of many kinds of cells, including neutrophils. To understand the function of CSF3R and recombinant human granulocyte colony-stimulating factor (rhCSF3) on melanocyte proliferation, this study compared the expression of CSF3R and the effects of rhCSF3 in primary human melanocytes, neutrophils and HEL 92.1.7 cells. The results show that CSF3R is localized in the cytoplasm and on cell membranes of melanocytes and neutrophils. The percentage of CSF3R(+) melanocytes was higher than CSF3R(+) HEL 92.1.7 cells, but was lower than CSF3R(+) neutrophils. Both CSF3R mRNA and CSF3R protein levels in melanocytes were higher than in HEL 92.1.7 cells, but were lower than in neutrophils. Treatment with rhCSF3 increased the proliferation of human melanocytes, but not their tyrosinase activity. Transcripts of CSF3R in human melanocytes, M14, A375 melanoma and A431 squamous cell carcinoma cells were also detected. Expression of the CSF3R V3 transcript was lower in melanocytes than in M14, A375 melanoma and A431 squamous cell carcinoma cells. In conclusion, rhCSF3 can promote melanocyte proliferation through CSF3R without affecting tyrosinase activity.

A comparative study of mitochondrial ultrastructure in melanocytes from perilesional vitiligo skin and perilesional halo nevi skin.

Vitiligo and halo nevi are both pigmentary disorders of the skin characterized by the acquired loss of functional epidermal melanocytes manifesting as white macules and patches. The cellular mechanism(s) and biochemical changes that result in the appearance of these two types of achromic lesions are still uncertain; and the relationship between vitiligo and halo nevi has been in dispute. In this study, we investigated the ultrastructure of mitochondria in melanocytes and in keratinocytes from perilesional vitiligo skin and from perilesional halo nevi skin using Transmission Electron Microscopy. Furthermore, we performed a quantitative analysis of mitochondrial morphology through a stereological study. As previously reported, we found that melanocytes from perilesional active vitiligo skin were loosely connected with their surroundings by their retracted dendrites. The surface density and the volume density of mitochondria in melanocytes and in keratinocytes from perilesional vitiligo skin are increased significantly compared with the controls, especially in active vitiligo. In contrast, there are no significant differences in mitochondria in melanocytes and in keratinocytes from perilesional halo nevi skin compared with the controls. In summary, the tendency of different morphologic alterations in mitochondria from perilesional vitiligo skin and from perilesional halo nevi skin reflect heterogeneous backgrounds between the two diseases, revealing that vitiligo and halo nevi may have separate pathogenic mechanisms. These findings may help elucidate the relationship of these two diseases and their underlying mechanisms.

The effects of calcipotriol on the dendritic morphology of human melanocytes under oxidative stress and a possible mechanism: is it a mitochondrial protector?

BACKGROUND: Vitiligo is an acquired pigmentary disorder of unknown etiology that is clinically characterized by the development of white macules in the skin related to the selective loss of melanocytes in those areas. Evidence shows that mitochondria might be a unifying target of reactive oxygen species (ROS) generation, cytokine production, catecholamine release and/or alteration of Ca(2+) metabolism that leads to melanocyte loss. OBJECTIVE: To assess the protective effect of calcipotriol on mitochondria of human melanocytes by investigating their dendritic morphology under oxidative stress. METHODS: Human melanocytes were treated with 0.05% H2O2 as well as various concentrations of calcipotriol, after which the retraction velocity of melanocyte dendrites was assessed. Detection of malondialdehyde (MDA) and superoxide dismutase (SOD) was performed as were the mitochondrial membrane potential (MMP) and intracellular calcium concentration ([Ca(2+)]i). Ultrastructural changes of mitochondria in melanocytes were observed by transmission electron microscopy. In addition, the expression of Beclin1, microtubule-associated protein 1 light chain 3 (LC3), dynamin related protein 1 (Drp1), mitofusin 1 (Mfn1) and mitofusin 2 (Mfn2), which are related to autophagy and mitochondrial dynamics, were analyzed by Western blot. RESULTS: Calcipotriol reduced the retraction velocity of melanocyte dendrites. In addition, calcipotriol, from 20nM to 80nM, decreased the level of MDA, increased the activity of SOD, suppressed the reduction of MMP and recovered Ca(2+) homeostasis by reducing [Ca(2+)]i in a concentration-dependent manner. Observation by transmission electron microscopy suggested that calcipotriol might reduce the injury of mitochondria in melanocytes under oxidative stress. Furthermore, the expression of Beclin1, LC3-II/LC3-I, Mfn2 and Drp1 was higher in the calcipotriol-treated melanocytes than in the control or H2O2-treated melanocytes. The level of Mfn1 was almost unchanged, but was higher at a concentration of 80nM calcipotriol than in any other condition. The expression of Mfn2 and Drp1 decreased with increasing calcipotriol concentration. CONCLUSION: Our study demonstrates the antioxidative effect of calcipotriol on melanocytes against oxidative damage. Moreover, calcipotriol could be a promising drug delivery strategy to protect melanocytes against oxidative damage in vitiligo through autophagy or mitophagy.

Stevens-Johnson Syndrome and toxic epidermal necrolysis: a multi-aspect comparative 7-year study from the People's Republic of China.

BACKGROUND: Stevens-Johnson Syndrome (SJS) and toxic epidermal necrolysis (TEN) are rare but severe cutaneous drug reactions. They are differentiated based on the fraction of the body surface area affected. Optimal therapy for SJS and TEN is a controversial issue. OBJECTIVE: We compared the treatments given to and the clinical outcomes of 39 cases of SJS and 48 cases of TEN seen at a single institution between January 2007 and December 2013 for better understanding of the clinical characteristics and development of the two conditions. METHODS: Demographic data, clinical characteristics, treatments given, and therapeutic responses observed were retrospectively collected. RESULTS: The incidence rates of hypoproteinemia and secondary infections are significantly higher in TEN than in SJS (P=0.001 and P=0.002, respectively). The corticosteroid dose did not influence the time from the initiation of therapy to control of the lesions in SJS, but increasing the dosage of corticosteroids progressively decreased the time from the initiation of therapy to control of the lesions in TEN. With increases in the utilization ratio of intravenous immunoglobulin (IVIG), the length of the hospital stay became shorter, whereas the time from the initiation of therapy to control of the lesions remained the same in SJS. However, for TEN, both the length of the hospital stay and the time from the initiation of therapy to control of the lesions became shorter with increases in the utilization ratio of IVIG. CONCLUSION: SJS and TEN are two variants of the same spectrum, and they differ from each other not only in the severity of epidermal detachment but also in other clinical parameters and their distinct clinical courses. Thus, differential treatment of both conditions may have benefits for their prognosis.

Fox gene loci in Takifugu rubripes and Tetraodon nigroviridis genomes and comparison with those of medaka and zebrafish genomes.

Members of the Fox gene family of transcriptional regulators are essential for animal development and have been extensively studied in vertebrates. The mouse and human genomes contain at least 40 FOX genes which are divided into 19 subclasses based on the sequence similarity of the highly conserved forkhead domain. Using the genome sequence of the Takifugu rubripes and Tetraodon nigroviridis , we examined the genomic complement of fox genes in these organisms to gain insight into the evolutionary relationship of this gene family. We identified 53 fox genes in Tetraodon nigroviridis and Takifugu rubripes genome by searching the forkhead domain. These genes are divided into 18 subclasses as follows: 8 fox genes in subclass O; 6 in subclass P ; 4 in subclasses D, J, and N; 3 in subclasses A, B, C, E, F, and I; 2 in subclasses K, L, and Q; and 1 in subclasses G, H, M, and R. Together with the forkhead domain sequences of human, chicken, frog, zebrafish, medaka, and Caenorhabditis elegans, the phylogenetic relationship of the fox genes in Takifugu rubripes and Tetraodon nigroviridis were analyzed and compared. The genes structure, general features, and the three-dimensional model of these genes were also discussed.

Structural, gene expression, and functional analysis of the fugu (Takifugu rubripes) insulin-like growth factor binding protein-4 gene.

The insulin-like growth factor (IGF) signaling pathway is a conserved pathway that regulates animal development, growth, metabolism, reproduction, and aging. The biological actions of IGFs are modulated by IGF-binding proteins (IGFBPs). Although the structure and function of fish IGFBP-1, -2, -3, and -5 have been elucidated, there is currently no report on the full-length structure of a fish IGFBP-4 nor its biological action. In this study, we cloned and characterized the IGFBP-4 gene from fugu. Sequence comparison, phylogenetic, and synteny analyses indicate that its chromosomal location, gene, and protein structure are similar to its mammalian orthologs. Fugu IGFBP-4 mRNA was easily detectable in all adult tissues examined with the exception of spleen. Older animals tended to have higher levels of IGFBP-4 mRNA in the muscle and eyes compared with younger animals. Starvation resulted in significant increases in IGFBP-4 mRNA abundance in the muscle, liver, gallbladder, and brain. Overexpression of fugu and human IGFBP-4 in zebrafish embryos caused a significant decrease in body size and somite number, suggesting that fugu IGFBP-4 inhibits growth and development, possibly by binding to IGFs and inhibiting their binding to the IGF receptors. These results provide new information about the structural and functional conservation, expression patterns, and physiological regulation of the IGFBP-4 gene in a teleost fish.

Transcript expression profiles of Takifugu rubripes spermatozoa and eggs by expressed sequence tag analysis.

Two cDNA libraries from Takifugu rubripes spermatozoa and eggs were constructed and a total of 620 expressed sequence tag (EST) clones were generated from the two libraries: 300 clones are from the spermatozoa library and 320 clones are from the eggs library. The most abundant cDNA clones in the two libraries were identified. A total of 207 'contigs' (or single) EST clones were found to share significant sequence identity with known sequences in the GenBank database, representing at least 51 different genes. In order to understand the two types of germ cells further, the expression profiles of the identified clones in these cDNA libraries were analyzed. Furthermore, the presence of specific messenger RNAs in the spermatozoa and eggs has been demonstrated with BLAST analysis; the spermatozoa and egg library can supply unique and novel cDNA sequences in the Takifugu rubripes EST project. Another aim of this study is to identify cDNA clones that can be used as molecular markers for the analysis of the spermatogenesis and oogenesis in Takifugu rubripes. Six potential clones (S1-3 from spermatozoa and E1-3 from eggs) were selected to analyze their expression patterns by reverse transcription (RT)-PCR analyses. Half of these showed a specific expression in the expected tissue. Two of the clones were found by RT-PCR and in situ hybridization to be expressed specifically in the testis or ovary, and they maybe suitable molecular markers for the analysis of spermatogenesis and oogenesis.

[Study on the safe edible of puffer fish resources].

In order to utilize the puffer fish resources furthest and to develop traditional diet culture, a project on the safe eating of puffer fish resources was conducted. The results showed that the distributing of teterodotoxin(TTX) in puffer body was not in uniformity. Owing that the content of TTX changes in different seasons and bodies, different methods were selected to crank out the delicious globefish food. The number of globefish food consumer has reached the amount to 121239 person-time since 1998. The technological process on teterodotoxin was proved reliable by mice bioassay and feeding trial in human.