Biomedical subjects

M Tran

Publications and source records attributed to M Tran.

At least 19 recordsLinked to original sources

PubMed2025

HLA-E and NKG2A Mediate Resistance to BCG Immunotherapy in Non-Muscle-Invasive Bladder Cancer.

Bacillus Calmette-Guérin (BCG) is the first-line therapy for high-grade non-muscle-invasive bladder cancer (NMIBC), yet many patients experience recurrence due to immune evasion. We identify HLA-E and NKG2A as mediators of adaptive resistance involving chronic activation of NK and T cells in BCG-unresponsive tumors. Prolonged IFN-γ exposure enhances HLA-E and PD-L1 expression on recurrent tumors, accompanied by the accumulation of NKG2A+ NK and CD8 T cells. HLA-Ehigh tumor cells preferentially cluster near CXCL12-rich stromal regions with dense effector cell presence, underscoring a spatially segregated tumor architecture. Although cytotoxic lymphocytes retain effector potential, their activity is restrained by HLA-E/NKG2A and PD-L1/PD-1 pathways located in their immediate neighborhood within the bladder tumor microenvironment. These data reveal a spatially organized immune escape program that limits anti-tumor immunity. Our findings support dually targeting NKG2A and PD-L1 checkpoint blockade as a rational, bladder-sparing strategy for patients with BCG-unresponsive NMIBC.

BCG-unresponsive↗

PubMed2006

Tissue culture and transient transformation of Marestail (Conyza canadensis (L.) Cronquist).

Glyphosate resistant crops are useful to agriculture by facilitating the use of nonselective herbicides, such as RoundUp, that have low human and environmental toxicity. The occurrence of glyphosate resistant weeds, however, has raised concern about the future utility of these crops. Conyza canadensis (L.) Cronquist (marestail or horseweed) is one such glyphosate resistant weed that has yet to be fully analyzed or established in tissue culture. Tissue culture enables the examination of physiological characteristics of a plant in an aseptic and controlled environment. For the present study, mairstail was cultured on a Murashige and Skoog based medium supplemented with 6-benzylaminopurine, and alpha-naphthaleneacetic acid. Plant regeneration was achieved on the same basal medium supplemented with only gibberellic acid. Glyphosate resistance could be demonstrated in the cultured tissues. The cultures could also be transformed with Agrobacterium tumefaciens without chemically inducing virulence using phenolics or glucose and resulting in a transformation frequency (transgenic events per total number of explants used) of about 13%. The tissue culture growth, preliminary glyphosate resistance data and genetic transformation data gathered in this project provide the means to further evaluate the mode of glyphosate resistance expressed by marestail.

Agrobacterium tumefaciens↗

PubMed2006

Transcriptome profiling of lung schistosomula,in vitro cultured schistosomula and adult Schistosoma japonicum.

The schistosomulum is the main target of vaccine-induced protective immunity; however, most studies have utilized schistosomula produced by mechanical transformation of infective larvae followed by in vitro culture rather than larvae isolated directly from the lungs of infected mammals. Using transmission electron microscopy, we demonstrated that there was little difference in the ultrastructure of Schistosoma japonicum schistosomula obtained by the two methods. However, significant differences in gene expression profiles were apparent when we used an oligonucleotide microarray to compare the gene expression profiles of schistosomula obtained in vivo from lung tissue with those maintained in vitro, and with adult worms of S. japonicum. It is likely that host environmental factors, which cannot be reliably reproduced in vitro, do influence the growth, development and overall biology of schistosomes. Thus caution is urged when using in vitro-cultured schistosomes and mechanically transformed/cultured schistosomula in molecular, biochemical and immunological studies.

Animals↗

PubMed2005

Factors that influence special care nursery admissions to a district hospital in South-western Sydney.

OBJECTIVES: To explore the socio-demographic factors and maternal characteristics that influence special care nursery (SCN) admission for infants of more than 34 weeks' gestation. Particularly, this paper aims (i) to estimate the incidence of SCN admission by various mothers' socio-demographic factors; and (ii) to investigate the relationship between SCN admission and mothers' socio-demographic and obstetric and gynaecological factors. METHODS: This was a cohort study of 10,148 pregnant women who accessed the birthing unit within a public district hospital in south-western Sydney in New South Wales, between 1998 and 2001. The main outcome measure was risk factors for SCN admission. RESULTS: The incidence of SCN admission was 11.7%. Multivariate analysis revealed that the risk factors for SCN admission were diabetes, gestational diabetes, high parity, pregnancy induced hypertension, living in suburbs with low education and occupation index, and no private health insurance status. CONCLUSIONS: The results from this large population-based study suggest that, apart from clinical/medical factors, admission to a special care nursery at a District Hospital was significantly affected by maternal insurance status and level of education and occupation.

Adult↗

PubMed2003

Risk factors for low birth weight in a socio-economically disadvantaged population: parity, marital status, ethnicity and cigarette smoking.

Low birth weight (LBW) is a public health problem, because it is associated with increased risk of morbidity and mortality. The principal aim of this study was to assess risk factors for LBW in a large multi-ethnic and socio-economically disadvantaged population. Data from 3242 mothers, who attended the Well Baby Clinic (Southwestern Sydney, Australia) for the first time, were analysed in relation to their demographic characteristics and socio-economic indices. The overall birthweight was 3377 +/- 577 g (mean +/- SD). In multiple linear regression analysis, smoking during pregnancy, marital status, parity, and country of birth were independently associated with birth weight. According to this analysis, lower birth weight was associated with mothers who had smoked during pregnancy (by 215.2 +/- 18.6 g), who were single (46.9 +/- 21 g), and of Asian background (108.5 +/- 38.2 g). However, higher parity was associated with significantly higher birth weight. The presence of each factor was coded as 1 and the absence, 0. A 'risk score' was then derived by summing up the individual scores. When birth weight was classified as 'low birth weight' (defined as those with birth weight being less than 2500 g) or normal birth weight, the overall prevalence of LBW was 1.9%. Each unit increase in the risk score was associated with a 1.9-fold (95% confidence interval: 1.5-2.6) increase in the risk of LBW. These data suggest that apart from marital status, ethnicity and parity, maternal smoking is the single most important preventable risk factors for LBW.

Female↗

PubMed2002

MUC1 epithelial mucin (CD227) is expressed by activated dendritic cells.

The MUC1 mucin (CD227) is a cell surface mucin originally thought to be restricted to epithelial tissues. We report that CD227 is expressed on human blood dendritic cells (DC) and monocyte-derived DC following in vitro activation. Freshly isolated murine splenic DC had very low levels of CD227; however, all DC expressed CD227 following in vitro culture. In the mouse spleen, CD227 was seen on clusters within the red pulp and surrounding the marginal zone in the white pulp. Additionally, we confirm CD227 expression by activated human T cells and show for the first time that the CD227 cytoplasmic domain is tyrosine-phosphorylated in activated T cells and DC and is associated with other phosphoproteins, indicating a role in signaling. The function of CD227 on DC and T cells requires further elucidation.

Animals↗

PubMed2001

Muc13, a novel human cell surface mucin expressed by epithelial and hemopoietic cells.

Transmembrane mucins are glycoproteins involved in barrier function in epithelial tissues. To identify novel transmembrane mucin genes, we performed a tblastn search of the GenBanktrade mark EST data bases with a serine/threonine-rich search string, and a rodent gene expressed in bone marrow was identified. We determined the cDNA sequence of the human orthologue of this gene, MUC13, which localizes to chromosome band 3q13.3 and generates 3.2-kilobase pair transcripts encoding a 512-amino acid protein comprised of an N-terminal mucin repeat domain, three epidermal growth factor-like sequences, a SEA module, a transmembrane domain, and a cytoplasmic tail (GenBanktrade mark accession no. ). MUC13 mRNA is expressed most highly in the large intestine and trachea, and at moderate levels in the kidney, small intestine, appendix, and stomach. In situ hybridization in murine tissues revealed expression in intestinal epithelial and lymphoid cells. Immunohistochemistry demonstrated the human MUC13 protein on the apical membrane of both columnar and goblet cells in the gastrointestinal tract, as well as within goblet cell thecae, indicative of secretion in addition to presence on the cell surface. MUC13 is cleaved, and the beta-subunit containing the cytoplasmic tail undergoes homodimerization. Including MUC13, there are at least five cell surface mucins expressed in the gastrointestinal tract.

Amino Acid Sequence↗

PubMed2001

Leber hereditary optic neuropathy, progressive visual loss, and multiple-sclerosis-like symptoms.

PURPOSE: To report a case of Leber hereditary optic neuropathy with multiple-sclerosis-like symptoms. METHODS: Observational case report. A 34-year-old man was found to have Leber hereditary optic neuropathy and a mutation at position 11778 of the mitochondrial genome. The progression of vision loss and onset of weakness in the right leg warranted neuroimaging. RESULTS: Magnetic resonance imaging documented multiple lesions in the brain and spinal cord. CONCLUSION: Although rarely reported, progression of optic neuropathy over months has been previously documented in Leber hereditary optic neuropathy. The emergence of multiple sclerosis-like symptoms and signs in our patient may be part of the spectrum of Leber hereditary optic neuropathy or a coincidental occurrence.

Adult↗

PubMed2001

Synthesis and SAR of 5-, 6-, 7- and 8-aza analogues of 3-aryl-4-hydroxyquinolin-2(1H)-one as NMDA/glycine site antagonists.

A series of 5-, 6-, 7- and 8-aza analogues of 3-aryl-4-hydroxyquinolin-2(1H)-one was synthesized and assayed as NMDA/glycine receptor antagonists. The in vitro potency of these antagonists was determined by displacement of the glycine site radioligand [(3)H]5,7-dicholorokynurenic acid ([(3)H]DCKA) in rat brain cortical membranes. Selected compounds were also tested for functional antagonism using electrophysiological assays in Xenopus oocytes expressing cloned NMDA receptor (NR) 1A/2C subunits. Among the 5-, 6-, 7-, and 8-aza-3-aryl-4-hydroxyquinoline-2(1H)-ones investigated, 5-aza-7-chloro-4-hydroxy-3-(3-phenoxyphenyl)quinolin-2-(1H)-one (13i) is the most potent antagonist, having an IC(50) value of 110 nM in [(3)H]DCKA binding and a K(b) of 11 nM in the electrophysiology assay. Compound 13i is also an active anticonvulsant when administered systemically in the mouse maximum electroshock-induced seizure test (ED(50)=2.3mg/kg, IP).

Animals↗

PubMed2001

Quality of health services and early postpartum discharge: results from a sample of non-English-speaking women.

Few studies have explored the knowledge, perceptions and satisfaction of ethnic women in relation to early obstetric discharge. The aim of this study was to form a profile of Vietnamese mothers who opt for early postpartum discharge, and to identify factors associated with those decisions. Focus group discussions, in-depth interviews and survey questionnaires were utilized to collect information from a sample of 160 mothers. Sixty mothers (38%) opted for early discharge (< 48 h). Factors strongly associated with early discharge were poor comprehension of English (OR=6.58, 95% CI=1.81, 24.02), very low level of education (OR=3.10, 95% CI=1.55, 6.20) and first time mother (OR=2.70, 95% CI=1.15, 4.27). The in-depth interviews identified further factors driving early discharge, including fear and anxiety when having to approach staff for assistance, and perceptions of disempowerment within culturally unfamiliar hospital surroundings. The findings in this study ought to alert administrators and clinicians to possible negative drivers for the choice of early postpartum discharge by Vietnamese mothers.

Adult↗

PubMed2001

[Standards, options and recommendations: Good clinical practice in the dietetic management of cancer patients: hospital catering].

CONTEXT: The "Standards, Options and Recommendations" (SOR) project, started in 1993, involves a collaboration between the Federation of the French Cancer Centres (FNCLCC), the 20 French Regional Cancer Centres, some French public university and general hospitals and private Clinics and medical scientific societies. Its main objective is the development of clinical practice guidelines to improve the quality of health care and outcome for cancer patients. The methodology is based on a literature review followed by a critical appraisal by a multidisciplinary group of experts to produce the draft guidelines which are then validated by specialists in cancer care delivery. OBJECTIVES: To develop clinical practice guidelines for hospital catering for cancer patient using the methodology developed by the Standards, Options and Recommendations project. METHODS: Data were identified by a literature search of Medline and the reference lists of experts in the groups. After the guidelines were drafted, they were validated by independent reviewers. RESULTS: The main recommendations are: 1) While taking into consideration the specific needs of cancer patients, the dietician is responsible for the hygiene, the sanitary quality of alimentation, the equilibrium and nutritional quality of the hospital catering. 2) Ordering and distribution of meals, and clearing up afterwards contribute to the quality of hospital catering and the personnel who do this should have time and be willing to listen to the patients. 3) The ordering of meals should be adapted to individual patient's requirements and must take into account the patient's medication. 4) The method of transporting the food chosen by the institution (cold or warm method) should be respected. The personnel responsible should receive regular and specific training to use the method correctly. 5) The intake of patients with nutritional follow-up should be reliably and reproducibly evaluated by the personnel after every meal. 6) Patient satisfaction should be assessed once a year and the results of this assessment used to improve the quality of hospital catering. 7) The dietician is the interface between the accounts department, the medical wards, the hospital catering department and the patients.

Adult↗

PubMed2000

Synthesis of N-substituted 4-(4-hydroxyphenyl)piperidines, 4-(4-hydroxybenzyl)piperidines, and (+/-)-3-(4-hydroxyphenyl)pyrrolidines: selective antagonists at the 1A/2B NMDA receptor subtype.

Antagonists at the 1A/2B subtype of the NMDA receptor (NR1A/2B) are typically small molecules that consist of a 4-benzyl- or a 4-phenylpiperidine with an omega-phenylalkyl substituent on the heterocyclic nitrogen. Many of these antagonists, for example ifenprodil (1), incorporate a 4-hydroxy substituent on the omega-phenyl group. In this study, the position of this 4-hydroxy substituent was transferred from the omega-phenyl group to the benzyl or phenyl group located on the 4-position of the piperidine ring. Analogues incorporating pyrrolidine in lieu of piperidine were also prepared. Electrical recordings using cloned receptors expressed in Xenopus oocytes show that high-potency antagonists at the NR1A/2B subtype are obtained employing N-(omega-phenylalkyl)-substituted 4-(4-hydroxyphenyl)piperidine, 4-(4-hydroxybenzyl)piperidine, and (+/-)-3-(4-hydroxyphenyl)pyrrolidine as exemplified by 21 (IC(50) = 0.022 microM), 33 (IC(50) = 0.059 microM), and 40 (IC(50) = 0.017 microM), respectively. These high-potency antagonists are >1000 times more potent at the NR1A/2B subtype than at either the NR1A/2A or NR1A/2C subtypes. The binding affinities of 21 at alpha(1)-adrenergic receptors ([(3)H]prazosin, IC(50) = 0.54 microM) and dopamine D2 receptors ([(3)H]raclopride, IC(50) = 1.2 microM) are reduced by incorporating a hydroxy group onto the 4-position of the piperidine ring and the beta-carbon of the N-alkyl spacer to give (+/-)-27: IC(50) NR1A/2B, 0.026; alpha(1), 14; D2, 105 microM. The high-potency phenolic antagonist 21 and its low-potency O-methylated analogue 18 are both potent anticonvulsants in a mouse maximal electroshock-induced seizure (MES) study (ED(50) (iv) = 0.23 and 0.56 mg/kg, respectively). These data indicate that such compounds penetrate the blood-brain barrier but their MES activity may not be related to NMDA receptor antagonism.

Animals↗

PubMed2000

Sequence of PHA synthase gene from two strains of Rhodospirillum rubrum and in vivo substrate specificity of four PHA synthases across two heterologous expression systems.

A 3.0-kb genomic fragment has been isolated from Rhodospirillum rubrum (ATCC 25903) that contains an open reading frame (ORF) with strong homology to other known polyhydroxyalkanoate (PHA) synthase genes. This ORF has lower homology to the R. rubrum strain Ha PHA synthase than would be expected within the same species. We have conducted a series of heterologous expression studies evaluating the in vivo substrate specificity of PHA synthase genes from Rhodobacter sphaeroides, Ralstonia eutropha (formerly Alcaligenes eutrophus), Thiocystis violacea, and Nocardia corrallina, within the PHA-synthase-negative hosts, Ralstonia eutropha DSM541 and Pseudomonas putida GpP104. The N. corrallina PHA synthase incorporated the highest percentage of C5 monomers in the polymer when fermented in medium supplemented with 0.1% heptanoate as the sole carbon source. When the T. violacea and R. sphaeroides were expressed in the PHA-negative host DSM541, a greater percentage of C5 monomer was observed in the polymer as compared to the expression of the PHA synthase of R. eutropha, when the transconjugants were fermented in medium supplemented with 0.4% propionate. Evaluation for preference of medium-chain-length monomers demonstrated the flexibility of the N. corrallina, T. violacea, and R. eutropha synthase genes to polymerize a copolyester composed of short- and medium-chain-length monomers when the respective transconjugants were fermented in medium supplemented with 0.5% octanoate. These studies demonstrate that the PHA synthase from N. corrallina, T. violacea, and R. eutropha are able to polymerize a copolyester composed of short- and medium-chain-length monomers, while the PHA synthase from R. sphaeroides lacks this ability and only produces a short-chain-length polymer. These observations suggest that the composition of the PHA from the PHA-producing organisms does not necessarily reflect the inherent specificity of the PHA synthase.

Acyltransferases↗

PubMed2000

Polyhydroxyalkanoate accumulation in Burkholderia sp.: a molecular approach to elucidate the genes involved in the formation of two homopolymers consisting of short-chain-length 3-hydroxyalkanoic acids.

Burkholderia sp. accumulates polyhydrox-yalkanoates (PHAs) containing 3-hydroxybutyrate and 3-hydroxy-4-pentenoic acid when grown on mineral media under limited phosphate or nitrogen, and using sucrose or gluconate as a carbon and energy source. Solvent fractionation and NMR spectroscopic characterization of these polyesters revealed the simultaneous accumulation of two homopolyesters rather than a co-polyester with random sequence distribution of the monomers [Valentin HE, Berger PA, Gruys KJ, Rodrigues MFA, Steinbuchel A, Tran M, Asrar J (1999) Macromolecules 32: 7389-7395]. To understand the genetic requirements for such unusual polyester accumulation, we probed total genomic DNA from Burkholderia sp. by Southern hybridization experiments using phaC-specific probes. These experiments indicated the presence of more than one PHA synthase gene within the genome of Burkholderia sp. However, when total genomic DNA from Burkholderia sp. was used to complement a PHA-negative mutant of Ralstonia eutropha for PHA accumulation, only one PHA synthase gene was obtained resembling the R. eutropha type of PHA synthases, based on amino acid sequence similarity. In addition to the PHA synthase gene, based on high sequence homology, genes encoding a beta-ketothiolase and acetoacetyl-CoA reductase were identified in a gene cluster with the PHA synthase gene. The arrangement of the three genes is quite similar to the R. eutropha poly-beta-hydroxybutyrate biosynthesis operon.

Acyltransferases↗

PubMed2000

Isolation, genomic organization, and expression analysis of the mouse and rat homologs of MEFV, the gene for familial mediterranean fever.

Familial Mediterranean fever (FMF) is a recessive disorder characterized by episodes of fever with serositis or synovitis. Recently the FMF gene (MEFV) was cloned; the protein product, pyrin/marenostrin, is thought to regulate inflammation in myeloid cells. In this manuscript we report the mouse and rat homologs of MEFV. The murine gene contains ten exons with a coding sequence of 2304 bp, while the rat homolog has nine exons with a coding sequence of 2253 bp. A considerable amino acid sequence homology was observed between the mouse and human (47.6% identity and 65.5% similarity) and between the mouse and rat genes (73.5% identity and 82.1% similarity). The predicted rodent proteins have several important domains and signals found in human pyrin, including a B-box zinc finger domain, Robbins-Dingwall nuclear localization signal, and coiled-coil domain. However, perhaps because of an ancient frame-shift mutation, neither the mouse nor the rat protein has an intact C-terminal B30.2 domain, in which most FMF-associated mutations have been found in human MEFV. Nevertheless, like the human gene, mouse Mefv is expressed in peripheral blood granulocytes but not lymphocytes. Consistent with its expression in granulocytes, Mefv was detected at high levels in the primary follicles and marginal zones of the splenic white pulp. Mefv is localized on mouse Chromosome (Chr) 16, region A3-B1, extending a region of synteny with human Chr 16p13.3. Development of knockout and knockin mouse models may provide further insights into the functional evolution of this gene.

Amino Acid Sequence↗

PubMed2000

In-situ evaluation of barrier-cream performance on human skin using laser-induced breakdown spectroscopy.

Laser-induced breakdown spectroscopy (LIBS) was used to evaluate the effect of barrier creams (skin protective creams) on human skin. A Nd: YAG laser at 1,064 nm was used with a pulse energy of 100 mJ. A method was developed to measure the effectiveness of barrier creams against zinc ion absorption from aqueous zinc chloride solution and oil paste zinc oxide, which represent model hydrophilic and lipophilic metal compounds, respectively. Zinc was chosen since it posed no risk to human skin. 3 representative commercial barrier creams advertised as being effective against lipophilic and hydrophilic substances were evaluated by measuring zinc absorbed through the stratum corneum. 4 consecutive skin surface biopsies (SSB) were taken from biceps of the forearms of 6 volunteers at time periods of 0.5 h and 3 h after application of the protective cream. Results were compared with control skin where no barrier cream was used. The zinc atomic emission line at 213.9 nm was selected. Gate delay and gate width time was optimized to obtain the best signal-to-noise ratio (SNR) and precision. This method provided a facile and rapid screening of the effectiveness of skin barrier creams against zinc ion penetration. The barrier creams were shown to provide appreciable protection against the penetration of both ZnCl2 and ZnO into the skin.

Adult↗

PubMed2000

Quality of life and home enteral tube feeding: a French prospective study in patients with head and neck or oesophageal cancer.

A prospective study was conducted to evaluate the impact of home enteral tube feeding on quality of life in 39 consecutive patients treated for head and neck or oesophageal cancer at the Centre François Baclesse in Caen, France. Patients were taken as their own controls. Quality of life was evaluated using the EORTC QLQ-C30 core questionnaire, and the EORTC H&N35 and OES24 specific questionnaires. The feeding technique tolerance was evaluated using a questionnaire specifically developed for this study. Two evaluations were made, the first a week after hospital discharge (n = 39) and the second 3 weeks later (n = 30). Overall, the global health status/quality of life scale score slightly improved; among symptoms, scale scores that significantly improved (P < 0.05) concerned constipation, coughing, social functioning and body image/sexuality. The physical feeding technique tolerance was acceptable while the technique was psychologically less tolerated with two-thirds of the patients longing to have the tube removed. One third of the patients was also uncomfortable about their body image. Home enteral tube feeding was responsible for not visiting family or close relations in 15% of patients, and not going out in public in 23%. We conclude that home enteral tube feeding is a physically well accepted technique although a substantial proportion of patients may experience psychosocial distress.

Adult↗

PubMed2000

Application of a propionyl coenzyme A synthetase for poly(3-hydroxypropionate-co-3-hydroxybutyrate) accumulation in recombinant Escherichia coli.

The genetic operon for propionic acid degradation in Salmonella enterica serovar Typhimurium contains an open reading frame designated prpE which encodes a propionyl coenzyme A (propionyl-CoA) synthetase (A. R. Horswill and J. C. Escalante-Semerena, Microbiology 145:1381-1388, 1999). In this paper we report the cloning of prpE by PCR, its overexpression in Escherichia coli, and the substrate specificity of the enzyme. When propionate was utilized as the substrate for PrpE, a K(m) of 50 microM and a specific activity of 120 micromol. min(-1). mg(-1) were found at the saturating substrate concentration. PrpE also activated acetate, 3-hydroxypropionate (3HP), and butyrate to their corresponding coenzyme A esters but did so much less efficiently than propionate. When prpE was coexpressed with the polyhydroxyalkanoate (PHA) biosynthetic genes from Ralstonia eutropha in recombinant E. coli, a PHA copolymer containing 3HP units accumulated when 3HP was supplied with the growth medium. To compare the utility of acyl-CoA synthetases to that of an acyl-CoA transferase for PHA production, PHA-producing recombinant strains were constructed to coexpress the PHA biosynthetic genes with prpE, with acoE (an acetyl-CoA synthetase gene from R. eutropha [H. Priefert and A. Steinbüchel, J. Bacteriol. 174:6590-6599, 1992]), or with orfZ (an acetyl-CoA:4-hydroxybutyrate-CoA transferase gene from Clostridium propionicum [H. E. Valentin, S. Reiser, and K. J. Gruys, Biotechnol. Bioeng. 67:291-299, 2000]). Of the three enzymes, PrpE and OrfZ enabled similar levels of 3HP incorporation into PHA, whereas AcoE was significantly less effective in this capacity.

Base Sequence↗