Biomedical subjects

Shan Lu

Publications and source records attributed to Shan Lu.

At least 19 recordsLinked to original sources

PubMed2025

Evolutionary legacy of the "living fossil" genus Parrotia (Hamamelidaceae): genomic insights into species divergence and polygenic adaptation.

Despite their long evolutionary history, the genomic basis of adaptation and speciation in "living fossil" plants remain largely unexplored. Parrotia, a Tertiary relict tree genus with two extant species, P. subaequalis and P. persica, exhibits a disjunct distribution between East Asia and West Asia. Here, we present the first chromosome-level assemblies for both species, confirmed their sibling relationship, and dated the speciation event to the early Miocene. The recent proliferation of long-terminal repeat retrotransposons has driven the genome expansion in P. subaequalis. We detected widespread heterogeneous genomic differentiation between species. Extensive signals of divergent selection, local adaptation, and elevated Ka/Ks ratios in Parrotia indicate that this genus has undergone adaptive evolution in distinct refugia, challenging the notion of it as an "evolutionary dead end". Our findings provide new insights into the genomic evolution, environmental adaptation, and speciation of this "living fossil" tree genus.

Genome, Plant↗

PubMed2025

Unveiling the inequivalent biochemical functions of OsSMO2-1 and OsSMO2-2 in rice phytosterol biosynthesis using a customized sterol standards mixture.

Phytosterols, natural active components widely distributed in plants, commonly include β-sitosterol, stigmasterol, and campesterol. However, identifying their synthesis intermediates can be challenging because access to corresponding commercial standard compounds is not always possible. To overcome these limitations, we synthesized phytosterol intermediates by co-infecting Nicotiana benthamiana with verified phytosterol synthases and identified them via gas chromatography-mass spectrometry analysis. Then, we mixed the identified intermediates, re-analyzed them by GC-MS to get a customized standard mixture, and established the corresponding standard spectrum library and characteristic ion database. To prove the usability of this standard mixture, we analyzed sterol intermediate types and contents in OsSMO2-1 and OsSMO2-2 rice mutants with this tool. Using the standard mixture, we analyzed sterolomics in wild-type rice buds at different germination days. As buds grew, detectable sterol intermediates decreased; only campesterol, sitosterol, and stigmasterol were found at the 9-day-old bud. When we compared mutants (ossmo2-1/ossmo2-2) with the wild type, we noted an obvious peak of 24-ethylidenelophenol accumulation in the ossmo2-1 mutant, indicating non-equivalent functions of OsSMO2-1 and OsSMO2-2 in sterol synthesis. OsSMO2-1 is more crucial for stigmasterol and sitosterol synthesis. This validates the standard mixture's utility.

Oryza↗

PubMed2020

Genomic Analysis of Circulating Tumor Cells at the Single-Cell Level.

Circulating tumor cells (CTCs) have a great potential for noninvasive diagnosis and real-time monitoring of cancer. A comprehensive evaluation of four whole genome amplification (WGA)/next-generation sequencing workflows for genomic analysis of single CTCs, including PCR-based (GenomePlex and Ampli1), multiple displacement amplification (Repli-g), and hybrid PCR- and multiple displacement amplification-based [multiple annealing and loop-based amplification cycling (MALBAC)] is reported herein. To demonstrate clinical utilities, copy number variations (CNVs) in single CTCs isolated from four patients with squamous non-small-cell lung cancer were profiled. Results indicate that MALBAC and Repli-g WGA have significantly broader genomic coverage compared with GenomePlex and Ampli1. Furthermore, MALBAC coupled with low-pass whole genome sequencing has better coverage breadth, uniformity, and reproducibility and is superior to Repli-g for genome-wide CNV profiling and detecting focal oncogenic amplifications. For mutation analysis, none of the WGA methods were found to achieve sufficient sensitivity and specificity by whole exome sequencing. Finally, profiling of single CTCs from patients with non-small-cell lung cancer revealed potentially clinically relevant CNVs. In conclusion, MALBAC WGA coupled with low-pass whole genome sequencing is a robust workflow for genome-wide CNV profiling at single-cell level and has great potential to be applied in clinical investigations. Nevertheless, data suggest that none of the evaluated single-cell sequencing workflows can reach sufficient sensitivity or specificity for mutation detection required for clinical applications.

Carcinoma, Non-Small-Cell Lung↗

PubMed2006

The cauliflower Or gene encodes a DnaJ cysteine-rich domain-containing protein that mediates high levels of beta-carotene accumulation.

Despite recent progress in our understanding of carotenogenesis in plants, the mechanisms that govern overall carotenoid accumulation remain largely unknown. The Orange (Or) gene mutation in cauliflower (Brassica oleracea var botrytis) confers the accumulation of high levels of beta-carotene in various tissues normally devoid of carotenoids. Using positional cloning, we isolated the gene representing Or and verified it by functional complementation in wild-type cauliflower. Or encodes a plastid-associated protein containing a DnaJ Cys-rich domain. The Or gene mutation is due to the insertion of a long terminal repeat retrotransposon in the Or allele. Or appears to be plant specific and is highly conserved among divergent plant species. Analyses of the gene, the gene product, and the cytological effects of the Or transgene suggest that the functional role of Or is associated with a cellular process that triggers the differentiation of proplastids or other noncolored plastids into chromoplasts for carotenoid accumulation. Moreover, we demonstrate that Or can be used as a novel genetic tool to induce carotenoid accumulation in a major staple food crop. We show here that controlling the formation of chromoplasts is an important mechanism by which carotenoid accumulation is regulated in plants.

Alleles↗

PubMed2006

Combination DNA plus protein HIV vaccines.

A major challenge in developing an HIV vaccine is to identify immunogens and delivery methods that will elicit balanced humoral and cell mediate immunities against primary isolates of HIV with diverse sequence variations. Since the discovery of using protein coding nucleic acids (mainly DNA but also possible RNA) as a means of immunization in the early 1990s, there has been rapid progress in the creative use of this novel approach for the development of HIV vaccines. Although the initial impetus of using DNA immunization was for the induction of strong cell-mediated immunity, recent studies have greatly expanded our understanding on the potential role of DNA immunization to elicit improved quality of antibody responses. This function is particularly important to the development of HIV vaccines due to the inability of almost every previous attempt to develop broadly reactive neutralizing antibodies against primary HIV-1 isolates. Similar to the efforts of developing cell mediated immunity by using a DNA prime plus viral vector boost approach, the best antibody responses with DNA immunization were achieved when a protein boost component was included as part of the immunization schedule. Current experience has suggested that a combination DNA plus protein vaccination strategy is able to utilize the benefits of DNA and protein vaccines to effectively induce both cell-mediated immunity and antibody responses against invading organisms.

Animals↗

PubMed2006

Ultrafast excited state dynamics of modified phthalocyanines: p-HPcZn and p-HPcCo.

Two modified metallophthalocyanines (MPcs) containing sulfonic naphthoxy substituents were synthesized. The measurements of transient absorption and time-resolved photoluminescence were used to study the ultrafast response and excited state dynamics of two MPcs in dimethyl sulfoxide (DMSO) solution, which were predominantly in the monomeric form. Under excitation at 400 nm, these molecules experience vibrational relaxation to the bottom of the first excited state and then the excitation rapidly converts to the low-lying charge-transfer (CT) state and finally reaches the triplet states. Under excitation at 800 nm, they show a two-photon absorption character, and their excited state dynamics exhibit strong dependence on the probe wavelength. The main results with 400 nm pumping are similar to the results with 800 nm pumping. For p-HPcZn, weak two-photon photoluminescence was also observed with a lifetime of 52 +/- 2 ps. A four-level model was used to illustrate the excited state dynamics of p-HPcZn, while a five-level model was suggested for p-HPcCo molecule.

Cobalt↗

PubMed2006

Hemagglutinin (HA) proteins from H1 and H3 serotypes of influenza A viruses require different antigen designs for the induction of optimal protective antibody responses as studied by codon-optimized HA DNA vaccines.

Effective antibody responses provide crucial immunity against influenza virus infection. The hemagglutinin (HA) protein is the major target of protective antibody responses induced by viral infection and by vaccination with both inactivated and live-attenuated flu vaccines, but knowledge about the optimal designs of protective HA antigens from different flu serotypes is still limited. In this study, we have significantly improved the immunogenicity of HA-expressing DNA vaccines by using codon-optimized HA sequences for either an H1 serotype (A/NewCal/20/99) or an H3 serotype (A/Panama/2007/99) human influenza A virus and then used these constructs as model antigens to identify the optimal HA antigen designs to elicit high-level protective antibody responses. Two forms of HA antigen, a wild-type, full-length HA and a secreted form with transmembrane (TM) domain-truncated HA, were produced. Both forms of HA DNA vaccines, from either H1 or H3 serotypes, were able to elicit high levels of HA-specific immunoglobulin G responses in immunized rabbits as measured by enzyme-linked immunosorbent assay. Interestingly, the abilities of H1 HA and H3 HA antigens to elicit hemagglutination inhibition (HI) and neutralizing antibody (NAb) responses differ. For the H1 HA antigens, the full-length HA induced significantly higher HI and NAb responses than did the TM-truncated HA. For the H3 HA antigen, both the full-length HA and TM-truncated HA induced high levels of HI and NAb responses. These data indicate that H1 and H3 antigens have different expression requirements for the induction of an optimal protective antibody response and that the structure integrity of HA antigens is critical for eliciting type-specific protective antibody responses. Our findings will have an important impact on future subunit-based flu vaccine development.

Animals↗

PubMed2006

Novel method for high-throughput phenotyping of sleep in mice.

Assessment of sleep in mice currently requires initial implantation of chronic electrodes for assessment of electroencephalogram (EEG) and electromyogram (EMG) followed by time to recover from surgery. Hence, it is not ideal for high-throughput screening. To address this deficiency, a method of assessment of sleep and wakefulness in mice has been developed based on assessment of activity/inactivity either by digital video analysis or by breaking infrared beams in the mouse cage. It is based on the algorithm that any episode of continuous inactivity of > or =40 s is predicted to be sleep. The method gives excellent agreement in C57BL/6J male mice with simultaneous assessment of sleep by EEG/EMG recording. The average agreement over 8,640 10-s epochs in 24 h is 92% (n = 7 mice) with agreement in individual mice being 88-94%. Average EEG/EMG determined sleep per 2-h interval across the day was 59.4 min. The estimated mean difference (bias) per 2-h interval between inactivity-defined sleep and EEG/EMG-defined sleep was only 1.0 min (95% confidence interval for mean bias -0.06 to +2.6 min). The standard deviation of differences (precision) was 7.5 min per 2-h interval with 95% limits of agreement ranging from -13.7 to +15.7 min. Although bias significantly varied by time of day (P = 0.0007), the magnitude of time-of-day differences was not large (average bias during lights on and lights off was +5.0 and -3.0 min per 2-h interval, respectively). This method has applications in chemical mutagenesis and for studies of molecular changes in brain with sleep/wakefulness.

Algorithms↗

PubMed2006

Immunogenicity and protection efficacy of monovalent and polyvalent poxvirus vaccines that include the D8 antigen.

Recent studies have established the feasibility of subunit-based experimental vaccines to protect animals from lethal poxvirus infection. Individual outer membrane proteins from intracellular and extracellular virions of vaccinia virus, when delivered in the form of either DNA vaccines or recombinant protein vaccines produced from baculovirus-infected insect cells, were able to protect mice from the vaccinia virus challenge and rhesus macaques from the monkeypox virus challenge. The polyvalent formulations with various combinations of the four poxvirus antigens (A27, L1, B5 and A33) achieved better protection than the monovalent formulation using only one of these antigens. However, it is not clear whether any of the remaining outer membrane poxvirus proteins can further improve the efficacy of the current polyvalent formulations. In this study, we conducted detailed analysis on the immunogenicity of D8, a previously reported protective antigen from intracellular mature virions. Our results indicated that D8 induced strong protective antibody responses and was effective in improving the efficacy of previously reported polyvalent poxvirus vaccine formulations. Therefore, D8 is an excellent candidate antigen to be included in the final polyvalent subunit-based poxvirus vaccines.

Animals↗

PubMed2006

Intracellular accumulation of a 46 kDa species of mouse prion protein as a result of loss of glycosylation in cultured mammalian cells.

Prion diseases are fatal neurodegenerative disorders characterized by the accumulation of an abnormal isoform (PrPSc) of the normal cellular prion protein (PrPC) in the brain. Reportedly, abnormal N-linked glycosylation patterns in PrPC are associated with disease susceptibility; thus, we compared the glycosylation status of normal and several mutant forms of the murine prion protein (MuPrP) in cultured mammalian cells. Substitution of the N-terminal signal sequence of normal MuPrP with a heterologous signal peptide did not alter glycosylation. When expressed without the C-terminal glycophosphatidylinositol anchor signal, the majority of MuPrP remained intracellular and unglycosylated, and a 46 kDa species (p46) of the unglycosylated PrPC was detected on reducing gels. p46 was also observed when wild-type MuPrP was expressed in the presence of tunicamycin or enzymatically deglycosylated in vitro. A rabbit polyclonal anti-serum raised against dimeric MuPrP cross-reacted with p46 and localized the signal within the Golgi apparatus. We propose that the 46 kDa signal is a dimeric form of MuPrP and in the light of recent studies, it can be argued that a relatively stable, non-glycosylated, cytoplasmic PrPC dimer, produced as a result of compromised glycosylation is an intermediate in initiating conversion of PrPC to PrPSc in sporadic transmissible spongiform encephalopathies.

Animals↗

PubMed2006

Characterization of TGF-beta-regulated interleukin-8 expression in human prostate cancer cells.

BACKGROUND: Interleukin (IL)-8 and transforming growth factor (TGF)-beta1 are overexpressed in advanced prostate cancer. The purpose of this study was to investigate TGF-beta1-regulated IL-8 expression in prostate cancer cells. METHODS: TGF-beta receptor expression was evaluated by real-time reverse-transcription PCR (RT-PCR) and Western blotting. TGF-beta1-regulated IL-8 expression was determined by real-time RT-PCR, enzyme-linked immunoabsorbance assay (ELISA), nuclear run-on, and IL-8 promoter reporter assay. RESULTS: PC-3MM2 cells expressed type I and type II TGF-beta receptors (TbetaRI and TbetaRII). LNCaP cells expressed significantly lower level of TbetaRII. Constitutive expression of IL-8 was detected in PC-3MM2 cells and LNCaP cells engineered with TbetaRII (LNCaP-TbetaRII). TGF-beta1 stimulated IL-8 expression in dose- and time-dependent manners, which was blocked by cycloheximide (CHX) and actinomycin D (ActD). The nuclear run-on and IL-8 luciferase reporter assays show that TGF-beta1 activated IL-8 gene transcription. CONCLUSIONS: TGF-beta1 signaling regulates IL-8 expression in prostate cancer cells and may contribute to the overexpression of IL-8 in human prostate cancer.

Activin Receptors, Type I↗

PubMed2006

Vav3 oncogene is overexpressed and regulates cell growth and androgen receptor activity in human prostate cancer.

The purpose of this research was to investigate the role of Vav3 oncogene in human prostate cancer. We found that expression of Vav3 was significantly elevated in androgen-independent LNCaP-AI cells in comparison with that in their androgen-dependent counterparts, LNCaP cells. Vav3 expression was also detected in other human prostate cancer cell lines (PC-3, DU145, and 22Rv1) and, by immunohistochemistry analysis, was detected in 32% (26 of 82) of surgical specimens of human prostate cancer. Knockdown expression of Vav3 by small interfering RNA inhibited growth of both androgen-dependent LNCaP and androgen-independent LNCaP-AI cells. In contrast, overexpression of Vav3 promoted androgen-independent growth of LNCaP cells induced by epidermal growth factor. Overexpression of Vav3 enhanced androgen receptor (AR) activity regardless of the presence or absence of androgen and stimulated the promoters of AR target genes. These effects of Vav3 could be attenuated by either phosphatidylinositol 3-kinase (PI3K) inhibitors or dominant-negative Akt and were enhanced by cotransfection of PI3K. Moreover, phosphorylation of Akt was elevated in LNCaP cells overexpressing Vav3, which could be blocked by PI3K inhibitors. Finally, we ascertained that the DH domain of Vav3 was responsible for activation of AR. Taken together, our data show that overexpression of Vav3, through the PI3K-Akt pathway, inappropriately activates AR signaling axis and stimulates cell growth in prostate cancer cells. These findings suggest that Vav3 overexpression may be involved in prostate cancer development and progression.

Blotting, Western↗

PubMed2006

Passive immunotherapy of Bacillus anthracis pulmonary infection in mice with antisera produced by DNA immunization.

Because of the high failure rate of antibiotic treatment in patients with anthrax there is a need for additional therapies such as passive immunization with therapeutic antibodies. In this study, we used codon-optimized plasmid DNAs (DNA vaccines) encoding Bacillus anthracis protective antigen (PA) to immunize rabbits for producing anti-anthrax antibodies for use in passive immunotherapy. The antisera generated with these DNA vaccines were of high titer as measured by ELISA. The antisera were also able to protect J774 macrophage cells by neutralizing the cytotoxic effect of exogenously added anthrax lethal toxin, and of the toxin released by B. anthracis (Sterne strain) spores following infection. In addition, the antisera passively protected mice against pulmonary challenge with an approximate 50 LD50 dose of B. anthracis (Sterne strain) spores. The protection in mice was obtained when the antiserum was given 1h before or 1h after challenge. We further demonstrated that IgG and F(ab')2 components purified from anti-PA rabbit hyperimmune sera retained similar levels of neutralizing activities against both exogenously added B. anthracis lethal toxin and toxin produced by B. anthracis (Sterne strain) spores. The high titer antisera we produced will enable an immunization strategy to supplement antibiotic therapy for improving the survival of patients with anthrax.

Animals↗

PubMed2006

Immunization of rhesus macaques with a polyvalent DNA prime/protein boost human immunodeficiency virus type 1 vaccine elicits protective antibody response against simian human immunodeficiency virus of R5 phenotype.

The immunogenicity of a poylvalent HIV-1 vaccine comprised of Env antigens from primary R5 isolates was evaluated in rhesus macaques. DNA vaccines encoding four Env antigens from multiple HIV-1 subtypes and HIV-1 Gag antigen from a single subtype elicited a persistent level of binding antibodies to gp120 from multiple HIV-1 isolates that were markedly enhanced following boosting with homologous gp120 proteins in QS-21 adjuvant irrespective of the route of DNA immunization. These sera neutralized homologous and, to a lesser degree, heterologous HIV-1 isolates. Four of the six immunized animals were completely protected following rectal challenge with a SHIV encoding Env from HIV-1(Ba-L), whereas the virus load was reduced in the remaining animals compared to naïve controls. Hence priming with DNA encoding Env antigens from multiple HIV-1 clades followed by boosting with homologous Env proteins elicits anti-HIV-1 immune responses capable of protecting macaques against mucosal transmission of R5 tropic SHIV isolate.

AIDS Vaccines↗

PubMed2006

Polyvalent HIV-1 Env vaccine formulations delivered by the DNA priming plus protein boosting approach are effective in generating neutralizing antibodies against primary human immunodeficiency virus type 1 isolates from subtypes A, B, C, D and E.

A major challenge in developing an HIV-1 vaccine is to identify immunogens and their delivery methods that can elicit broad neutralizing antibodies against primary isolates of different genetic subtypes. Recently, we demonstrated that priming with DNA vaccines expressing primary HIV-1 envelope glycoprotein (Env) followed by recombinant Env protein boosting was successful in generating positive neutralizing antibody responses against a clade B primary HIV-1 isolate, JR-FL, that was not easily neutralized. In the current study, we examined whether the DNA priming plus recombinant protein boosting approach delivering a polyvalent primary Env formulation was able to generate neutralizing antibodies against primary HIV-1 viral isolates from various genetic subtypes. New Zealand White rabbits were first immunized with DNA vaccines expressing one, three or eight primary HIV-1 gp120 antigens delivered by a gene gun followed by recombinant gp120 protein boosting. Neutralizing antibody responses were examined by two independently executed neutralization assays: the first one was a single round infection neutralization assay against a panel of 10 primary HIV-1 isolates of subtypes A, B, C and E and the second one used the PhenoSense assay against a panel of 12 pseudovirues expressing primary HIV-1 Env antigens from subtypes A, B, C, D and E as well as 2 pseudoviruses expressing the Env antigens from MN and NL4-3 viruses. Rabbit sera immunized with the DNA priming plus protein boosting approach, but not DNA vaccine alone or Env protein alone, were capable of neutralizing 7 of 10 viruses in the first assay and 12 of 14 viruses in the second assay. More importantly, sera immunized with the polyvalent Env antigens were able to neutralize a significantly higher percentage of viruses than the sera immunized with the monovalent antigens. Our results suggest that DNA priming followed by recombinant Env protein boosting can be used to deliver polyvalent Env-antigen-based HIV-1 vaccines to elicit neutralizing antibody responses against viruses with diverse genetic sequence variations.

AIDS Vaccines↗

PubMed2006

Experimental synchronization of independent entangled photon sources.

We report the generation of independent entangled photon pairs from two synchronized but mutually incoherent laser sources. The quality of synchronization is confirmed by observing a violation of Bell's inequality with 3.2 standard deviations in an entanglement swapping experiment. The techniques developed in our experiment are not only important for realistic linear optical quantum-information processing, but also enable new tests of local realism.

Journal Article↗

PubMed2006

Cyclin D1b variant influences prostate cancer growth through aberrant androgen receptor regulation.

Cyclin D1 is a multifaceted regulator of both transcription and cell-cycle progression that exists in two distinct isoforms, cyclin D1a and D1b. In the prostate, cyclin D1a acts through discrete mechanisms to negatively regulate androgen receptor (AR) activity and thus limit androgen-dependent proliferation. Accordingly, cyclin D1a is rarely overexpressed in prostatic adenocarcinoma and holds little prognostic value in this tumor type. However, a common polymorphism (A870) known to facilitate production of cyclin D1b is associated with increased prostate cancer risk. Here we show that cyclin D1b is expressed at high frequency in prostate cancer and is up-regulated in neoplastic disease. Furthermore, our data demonstrate that, although cyclin D1b retains AR association, it is selectively compromised for AR regulation. The altered ability of cyclin D1b to regulate the AR was observed by using both in vitro and in vivo assays and was associated with compromised regulation of AR-dependent proliferation. Consistent with previous reports, expression of cyclin D1a inhibited cell-cycle progression in AR-dependent prostate cancer cells. Strikingly, cyclin D1b significantly stimulated proliferation in this cell type. AR-negative prostate cancer cells were nonresponsive to cyclin D1 (a or b) expression, indicating that defects in AR corepressor function yield a growth advantage specifically in AR-dependent cells. In summary, these studies indicate that the altered AR regulatory capacity of cyclin D1b contributes to its association with increased prostate cancer risk and provide evidence of cyclin D1b-mediated transcriptional regulation.

Androgens↗

PubMed2006

beta-Carotene accumulation induced by the cauliflower Or gene is not due to an increased capacity of biosynthesis.

The cauliflower (Brassica oleracea L. var. botrytis) Or gene is a rare carotenoid gene mutation that confers a high level of beta-carotene accumulation in various tissues of the plant, turning them orange. To investigate the biochemical basis of Or-induced carotenogenesis, we examined the carotenoid biosynthesis by evaluating phytoene accumulation in the presence of norflurazon, an effective inhibitor of phytoene desaturase. Calli were generated from young seedlings of wild type and Or mutant plants. While the calli derived from wild type seedlings showed a pale green color, the calli derived from Or seedlings exhibited intense orange color, showing the Or mutant phenotype. Concomitantly, the Or calli accumulated significantly more carotenoids than the wild type controls. Upon treatment with norflurazon, both the wild type and Or calli synthesized significant amounts of phytoene. The phytoene accumulated at comparable levels and no major differences in carotenogenic gene expression were observed between the wild type and Or calli. These results suggest that Or-induced beta-carotene accumulation does not result from an increased capacity of carotenoid biosynthesis.

Brassica↗