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Biomedical subjects

Arun Sharma

Publications and source records attributed to Arun Sharma.

At least 19 recordsLinked to original sources

NeuroD and reaggregation induce beta-cell specific gene expression in cultured hepatocytes.

BACKGROUND: Our goal was to convert adult mouse hepatocytes to pancreatic beta-cells. METHODS AND RESULTS: To facilitate conversion, cultured primary hepatocytes were dedifferentiated by the removal of dexamethasone (Dex) from the culture media. Removal of Dex caused detachment of hepatocytes from the culture dish, but the addition of betacellulin prevented this from happening. With the combination of lack of Dex and addition of betacellulin, albumin mRNA levels decreased. Cultured hepatocytes had a faint expression of insulin 2 mRNA, Nkx 6.1 and Pax 6 mRNA. Dedifferentiated hepatocytes were transduced with adenoviruses expressing NeuroD1, Ngn 3, or Pax 4. NeuroD1 transduction increased the insulin 2 mRNA but caused detachment of cells. However, when hepatocytes were allowed to reaggregate for 4 and 6 days in hydrophobic plates after transduction with NeuroD1, further increases of insulin 2 mRNA were found along with induction of PDX-1, IAPP, NeuroD1, Ngn3, Pax 4, Isl-1, PC1, PC2 and islet glucokinase. Additionally, glucagon, pancreatic polypeptide and somatostatin expression were induced, but neither elastase 1 nor insulin 1 mRNA could be detected. Ngn 3 and Pax 4 had effects similar to NeuroD1, but did not increase insulin 2 mRNA as much as NeuroD1. CONCLUSION: We conclude that the combination of NeuroD1 and reaggregation promotes cultured dedifferentiated hepatocytes to differentiate towards a pancreatic beta-cell phenotype.

Animals↗

A switch from MafB to MafA expression accompanies differentiation to pancreatic beta-cells.

Major insulin gene transcription factors, such as PDX-1 or NeuroD1, have equally important roles in pancreatic development and the differentiation of pancreatic endocrine cells. Previously, we identified and cloned another critical insulin gene transcription factor MafA (RIPE3b1) and reported that other Maf factors were expressed in pancreatic endocrine cells. Maf factors are important regulators of cellular differentiation; to understand their role in differentiation of pancreatic endocrine cells, we analyzed the expression pattern of large-Maf factors in the pancreas of embryonic and adult mice. Ectopically expressed large-Maf factors, MafA, MafB, or cMaf, induced expression from insulin and glucagon reporter constructs, demonstrating a redundancy in their function. Yet in adult pancreas, cMaf was expressed in both alpha- and beta-cells, and MafA and MafB showed selective expression in the beta- and alpha-cells, respectively. Interestingly, during embryonic development, a significant proportion of MafB-expressing cells also expressed insulin. In embryos, MafB is expressed before MafA, and our results suggest that the differentiation of beta-cells proceeds through a MafB+ MafA- Ins+ intermediate cell to MafB- MafA+ Ins+ cells. Furthermore, the MafB to MafA transition follows induction of PDX-1 expression (Pdx-1(high)) in MafB+ Ins+ cells. We suggest that MafB may have a dual role in regulating embryonic differentiation of both beta- and alpha-cells while MafA may regulate replication/survival and function of beta-cells after birth. Thus, this redundancy in the function and expression of the large-Maf factors may explain the normal islet morphology observed in the MafA knockout mice at birth.

Animals↗

Estimation of aroma glycosides of nutmeg and their changes during radiation processing.

Glycosidically bound volatile compounds of nutmeg were identified as glyco-conjugates of p-cymene-7-ol, eugenol, methoxyeugenol and alpha-terpineol. Using phenyl-beta-glucoside as external standard the contents of these glycosidic precursors were estimated based on the measurement of TLC spot density on a densitometer. p-Cymene-7-ol rutinoside was the major aroma glycoside (3.15 mg/100 g), followed by glucosides of methoxyeugenol (0.61 mg/100 g), eugenol (0.50 mg/100 g) and alpha-terpineol (0.51 mg/100 g). A dose-dependent breakdown of these glycosidic precursors was observed during gamma-radiation processing. Among the four glycosides, alpha-terpineol glucoside was the most sensitive to radiation while p-cymene-7-ol rutinoside was the least sensitive. A reduction in the content of total glycosides by almost 50% was noted at a dose of 5kGy. Partitioning of aroma glycoside into n-butanol from aqueous extracts was found to result in rapid isolation of aroma glycosides, avoiding time consuming pre-purification on Amberlite XAD-2 column. A routine method based on extraction into n-butanol and subsequent quantification of post-irradiation changes in aroma glycosides on a TLC plate using a densitometer is proposed.

Chromatography, Thin Layer↗

Complications associated with fixed partial dentures with a loose retainer.

STATEMENT OF PROBLEM: When 1 retainer of a definitive fixed partial denture (FPD) becomes loose, the clinician has the option of either sectioning and removing the FPD or attempting removal of the intact cemented retainer. Excessive force during attempted FPD removal may irreversibly damage dentition. PURPOSE: This study evaluated the type and frequency of complications that may accompany the removal of definitive cemented FPDs with a single loose retainer. MATERIAL AND METHODS: Participants included 22 patients with a definitive cemented FPD with complete cast crown coverage castings in which 1 retainer became loose and the other retainer remained cemented. Cemented FPD retainers were removed using 1 of 2 attachments supplied with a crown removal system (Dentco). The patient and FPD characteristics, type of attachment, number of attempts before removal, condition of removed FPD, and condition of the abutment teeth were recorded. Descriptive statistics were used to analyze the data. RESULTS: Clinical findings recorded prior to FPD removal showed that 41% of patients were unaware they had a loose FPD retainer and 82% reported no discomfort associated with the loose retainer. Caries were noted on 50% of the teeth with a loose retainer. Damage resulting from attempted removal of the cemented retainer included minor porcelain fracture (9%), minor core chipping (14%), minor incisal edge chipping of tooth preparations (27%), and major damage to the abutment tooth (4%). CONCLUSION: Retrieval of an intact FPD and recementation was possible 64% of the time.

Adult↗

Molecules involved in the modulation of rapid cell death in Xanthomonas.

In earlier studies from this laboratory, Xanthomonas campestris pv. glycines was found to exhibit a nutrition stress-related postexponential rapid cell death (RCD). The RCD was exhibited in protein-rich media but not in starch or other minimal media. This RCD in X. campestris pv. glycines was found to display features similar to those of the programmed cell death (PCD) of eukaryotes. Results of the present study showed that the observed RCD in this organism is both positively and negatively regulated by small molecules. The amino acids glycine and l-alanine as well as the D isomers of valine, methionine, and threonine were found to induce the synthesis of an active caspase-3-like protein that was associated with the onset of RCD. Addition of pyruvate and citrate to the culture medium induced both the synthesis of active caspase-3-like protein and RCD. Higher levels of intracellular accumulation of pyruvate and citrate were also observed under conditions favoring RCD. On the other hand, dextrin and maltose, the hydrolytic products of starch, inhibited the synthesis of the caspase-3-like protein. Addition of glucose and cyclic AMP (cAMP) to the RCD-favoring medium prevented RCD. Glucose, cAMP, caffeine (a known inhibitor of a phosphodiesterase that breaks down cAMP), and forskolin (from the herb Coleus forskholii, known to activate the enzyme adenylate cyclase that forms cAMP) inhibited the caspase enzyme activity in vivo and consequently the RCD process. The addition of glucose and other inhibitors of RCD enhanced intracellular cAMP accumulation. This is the first report demonstrating the involvement of small molecules in the regulation of nutrition stress-related stationary-phase rapid cell death in X. campestris pv. glycines, which is programmed.

Amidohydrolases↗

Evidence for a role of the ubiquitin-proteasome pathway in pancreatic islets.

The ubiquitin-proteasome pathway is crucial for protein turnover. Part of the pathway involves deubiquitination, which is carried out by cystein proteases known as ubiquitin COOH-terminal hydrolases. The isoform Uch-L1 was found to be present in large amounts in rat islets by immunostaining, Western blot analysis, and RT-PCR. Culturing islets in high glucose concentrations (16.7 mmol/l) for 24 h led to decreased gene expression. Exposure to chronic hyperglycemia following 90% partial pancreatectomy also led to reduced Uch-L1 expression. Expression of other members of the ubiquitin-proteasome pathway studied after culturing islets at high glucose concentrations revealed little change except for modest declines in parkin, human ubiquitin-conjugating enzyme 5 (UbcH5), and beta-TRCP (transducin repeat-containing protein). With the pancreatectomy model, expression of polyubiquitin-B and c-Cbl were increased and E6-associated protein was reduced. Further insight about the proteasome pathway was obtained with the proteasome inhibitor lactacystin, which in short-term 2-h experiments enhanced glucose-induced insulin secretion. An important role for the ubiquitin-proteasome pathways in beta-cells is suggested by the findings that changes in glucose concentration influence expression of genes in the pathway and that blockade of the proteasome degradation machinery enhances glucose-stimulated insulin secretion.

Base Sequence↗

Microbiological evaluation of sprouts marketed in Mumbai, India, and its suburbs.

A study was undertaken to assess the microbiological quality of sprouts marketed in Mumbai and its suburbs. A total of 124 sprout samples of four different legumes--mung (Phaseolus aureus), matki (Phaseolus aconitifolius), chana (Cicer arietinum), and vatana (Pisum sativum)--were analyzed over a period of 12 months for aerobic plate counts, coliforms, yeast and mold counts, staphylococci, Salmonella, Listeria monocytogenes, Escherichia coli, E. coli O157:H7, and coagulase-positive Staphylococcus aureus. Aerobic plate counts ranged from 7.6 to 8.9 log CFU/g, coliform counts ranged from 5.4 to 7.9 log CFU/g, yeast and mold counts ranged from 3.6 to 7.3 log CFU/g, and staphylococci counts ranged from 3.3 to 6.6 log CFU/ g. Nonpathogenic E. coli was detected in 13% of the mung, 26% of the matki, 40% of the chana, and 19% of the vatana samples. Salmonella Typhimurium was detected in 21% of the mung, 40% of the matki, and 4% of the chana samples. Salmonella Dublin was detected in 2% of the mung samples, and Salmonella Washington was detected in 4% of the matki samples. L. monocytogenes and E. coli O157:H7 were not detected in any of the samples examined. Coagulase-positive S. aureus was detected in 4% of the mung, 11% of the matki, and 4% of the chana samples. The results indicated that the marketed sprouts were of poor microbiological quality; therefore, further processing, such as radiation processing, is needed to ensure their safety.

Bacteria↗

Effectiveness of radiation processing in elimination of Salmonella typhimurium and Listeria monocytogenes from sprouts.

The effectiveness of radiation treatment in eliminating Salmonella Typhimurium and Listeria monocytogenes on laboratory inoculated ready-to-eat sprouts was studied. Decimal reduction doses (D10-values) for Salmonella Typhimurium and L. monocytogenes in dry seeds of mung (green gram), matki (dew gram), chana (chick pea), and vatana (garden pea) ranged from 0.189 to 0.303 kGy and 0.294 to 0.344 kGy, respectively. In sprouts made from these seeds, the D10-values ranged from 0.192 to 0.208 kGy for Salmonella Typhimurium and from 0.526 to 0.588 kGy for L. monocytogenes. Radiation treatment with a 2-kGy dose resulted in complete elimination of 10(4) CFU/g of Salmonella Typhimurium and 10(3) CFU/g of L. monocytogenes from all the four varieties of sprouts. No recovery of Salmonella Typhimurium and L. monocytogenes was observed in the radiation treated samples stored at 4 and 8 degrees C up to 12 days. Radiation treatment with 1 kGy and 2 kGy resulted in a reduction of aerobic plate counts and coliform counts by 2 and 4 log CFU/g, respectively; the yeast and mold counts and staphylococci counts decreased by 1 and 2 log CFU/g, respectively. However, during postirradiation storage at 4 and 8 degrees C, aerobic plate counts, coliform counts, yeast and mold counts, and staphylococci counts remained constant throughout the incubation period. This study demonstrates that a 2-kGy dose of irradiation could be an effective method of processing to ensure microbial safety of sprouts.

Colony Count, Microbial↗

A pragmatic, three-arm randomised controlled trial of spiritual healing for asthma in primary care.

BACKGROUND: Well-designed trials are required to assess if complementary and alternative medicine (CAM) is effective. AIM: This study assessed the effectiveness of spiritual healing for asthma. DESIGN OF STUDY: Randomised, placebo-controlled trial. SETTING: Aberdeen, Scotland. METHOD: This was a single-blind, three-armed randomised, controlled trial of spiritual healing for asthma, comparing the effectiveness of five sessions of spiritual healing with placebo (delivered by an actor), and with a control group receiving normal care only. The primary outcome measure was the Juniper Asthma Quality of Life Questionnaire (AQLQ). Secondary outcomes were forced expiratory flow in one second (FEV1), peak expiratory flow (PEF), HADS (Hospital Anxiety Depression Scale), SF-36 and MYMOP (Measure Yourself Medical Outcome Profile). Baseline and follow-up data were collected. RESULTS: Eighty-eight adult patients receiving pharmacological treatment for asthma participated. AQLQ scores improved significantly from baseline and the end of treatment in all groups (spiritual healing P = 0.008; 'sham' healing P = 0.001 and control P = 0.01) but there was no significant difference between groups (P = 0.57). These improvements were maintained at follow-up 1 for two of the groups (spiritual healing P = 0.016; sham healing P = 0.001 and control P = 0.09) but none of the groups showed an improvement at follow-up 2 (spiritual healing P = 0.161; sham healing P = 0.016 and control P = 0.11). Similar proportions of patients in each group showed a clinically important improvement in AQLQ score. Analysis of AQLQ scores at end of treatment and both follow-up periods indicated no significance between group differences. No consistent changes were seen in secondary outcome measures, possibly due to the small sample size. CONCLUSION: Spiritual healing does not appear to have any specific affect on patient asthma related quality of life.

Adult↗

Regulation of insulin gene expression by overlapping DNA-binding elements.

The transcription factor MafA/RIPE3b1 is an important regulator of insulin gene expression. MafA binds to the insulin enhancer element RIPE3b (C1-A2), now designated as insulin MARE (Maf response element). The insulin MARE element shares an overlapping DNA-binding region with another insulin enhancer element A2. A2.2, a beta-cell-specific activator, like the MARE-binding factor MafA, binds to the overlapping A2 element. Our previous results demonstrated that two nucleotides in the overlapping region are required for the binding of both factors. Surprisingly, instead of interfering with each other's binding activity, the MafA and the A2-binding factors co-operatively activated insulin gene expression. To understand the molecular mechanisms responsible for this functional co-operation, we have determined the nucleotides essential for the binding of the A2.2 factor. Using this information, we have constructed non-overlapping DNA-binding elements and their derivatives, and subsequently analysed the effect of these modifications on insulin gene expression. Our results demonstrate that the overlapping binding site is essential for maximal insulin gene expression. Furthermore, the overlapping organization is critical for MafA-mediated transcriptional activation, but has a minor effect on the activity of A2-binding factors. Interestingly, the binding affinities of both MafA and A2.2 to the overlapping or non-overlapping binding sites were not significantly different, implying that the overlapping binding organization may increase the activation potential of MafA by physical/functional interactions with A2-binding factors. Thus our results demonstrate a novel mechanism for the regulation of MafA activity, and in turn beta-cell function, by altering expression and/or binding of the A2.2 factor. Our results further suggest that the major downstream targets of MafA will in addition to the MARE element have a binding site for the A2.2 factor.

Animals↗

Tympanoplasty type I in children--an evaluative study.

OBJECTIVE: A considerable controversy surrounds the subject of paediatric tympanoplasty type I. Conflicting views regarding timing of surgery and various other factors like Eustachian tube, status of contralateral ear, site and type of perforation etc. are presented by various published case series in medical literature. However, very few prospective studies address the issue. Thus, a prospective study was planned to evaluate the success rate of paediatric tympanoplasty type I and analyse the factors suggested to influence the outcome of the said surgery. METHOD: A cohort study with control was carried out in 40 patients of chronic suppurative otitis media-mucosal disease in a tertiary care teaching hospital. Patients were divided into two groups of 20 cases each, one comprising of paediatric patients (8-14 years) and other comprising of adult patients (more than 14 years of age). All the patients underwent tympanoplasty type I by post auricular inlay technique. An intact graft at the end of 6th month was considered a success and a minimum hearing improvement of 10 db in two consecutive frequencies was regarded as an audiological success. Results were tabulated and statistically analysed using chi2-test (with Yates correction). RESULTS: A success rate of 80% in terms of graft uptake and hearing improvement of 61% was seen in paediatric cases, compared to 85% graft uptake and 65% hearing improvement in adult cases. Statistically, on applying chi2-test, the value of P is >0.05 which is insignificant was obtained, indicating that age does not influence paediatric tympanoplasty type I. Also, the data for evaluating the correlation between Eustachian tube and paediatric tympanoplasty type I showed no statistical significance. In addition, no association between site and type of perforation and paediatric tympanoplasty type I was observed when results were compared to adult patients. However, anteriorly placed perforations were found to give poor results in both adult and paediatric group. CONCLUSIONS: The results of this cohort study mitigates against delaying tympanoplasty type I in paediatric population. This study also elucidates that Eustachian tube, contralateral ear pathology, site and type of perforation do not influence the outcome of the said surgery. However, anteriorly placed perforations require vigilance while repair in view of high failure rate. We recommend the age of 8 years for the paediatric tympanoplasty type I, but in view of redundancy of eustachian tube factor, the criteria for delineating minimum age for success of the said surgery is open to debate. More prospective studies are required in this direction.

Adolescent↗

Potato peel extract-a natural antioxidant for retarding lipid peroxidation in radiation processed lamb meat.

The effective utilization of potato peel, a waste generated in large quantities by the food industry, as an antioxidant was investigated. Potato peel extract (PPE) exhibited high phenolic content (70.82 mg of catechin equivalent/100 g), chlorogenic acid (27.56 mg/100 g of sample) being the major component. The yield of total phenolics and chlorogenic acid increased by 26 and 60%, respectively, when the extract was prepared from gamma irradiated (150 Gy) potatoes. PPE showed excellent antioxidant activity as determined by beta-carotene bleaching and radical scavenging activity of 1,1-diphenyl-2-picrylhydrazyl (DPPH). The suitability of PPE for controlling lipid oxidation of radiation processed lamb meat was also investigated. PPE (0.04%) when added to meat before radiation processing was found to retard lipid peroxidation of irradiated meat as measured by TBA number and carbonyl content. The antioxidant activity of PPE was found to be comparable to butylated hydroxytoluene (BHT).

Animals↗

The effects of Omega-3 fatty acids on growth regulation of epithelial ovarian cancer cell lines.

OBJECTIVE: Omega-3 fatty acids (OM-3FAs) have been shown to possess anti-carcinogenic properties. We investigated the effect of OM-3FAs on epithelial ovarian cancer cell lines to determine if a growth suppressive effect existed and to gain a better insight on the potential molecular mechanisms that may be involved. METHODS: Three epithelial ovarian cancer cell lines (SKOV-3 [p53 null], TOV-21G [wt p53] and OVCAR-3 [mutant p53]) and one immortalized ovarian surface epithelial cell line (IOSE-29 [wt p53]) were treated with OM-3FAs and evaluated for cellular proliferation (WST-1 assay), apoptosis (Annexin V-FITC/PI method) and VEGF expression (VEGF ELISA assay). RESULTS: A statistically significant inhibitory effect under the influence of OM-3FAs was detected in all four cell lines. Apoptosis and VEGF down-regulation were either limited or not detected in the p53 null and mutant cell lines, SKOV-3 and OVCAR-3 respectively. Apoptosis and/or VEGF down-regulation was strongly evident in the wt p53 cell lines TOV-21G and IOSE-29. CONCLUSION: These data suggest that, under the influence of OM-3FAs, there are definitive growth suppressive mechanisms at work and that the biologic effects of OM-3FAs may in part be mediated by the p53 status.

Apoptosis↗

Purification and characterization of a hemoglobin degrading aspartic protease from the malarial parasite Plasmodium vivax.

Aspartic proteases of human malarial parasites are thought to play key roles in essential pathways of merozoite release, invasion and host cell hemoglobin degradation during the intraerythrocytic stages of their life cycle. Therefore, we have purified and characterized Plasmodium vivax aspartic protease, to determine if this enzyme can be used as potential drug target/immunogen, and its inhibitors as potential antimalarial drug. The P. vivax aspartic protease has been purified by a combination of ion exchange and size exclusion chromatographies and HPLC. Its properties were examined in order to define a role in the hemoglobin degradation process. The purified enzyme migrated as a single band on native PAGE and SDS/PAGE with a molecular mass of 40 kDa. Gelatin zymogram analyses revealed a clear zone of proteolytic activity corresponding to the band obtained on native PAGE and SDS/PAGE. The enzyme has an optimal pH of 4.0 and exhibits its highest activity at 37 degrees C. The enzyme is inhibited by pepstatin, but not by other inhibitors including o-phenanthroline, EDTA, PMSF or E-64, supporting its designation as an aspartic protease; its IC50 value was found to be 3.0 microM. A Lineweaver Burk double reciprocal plot with pepstatin shows that the inhibition is competitive with respect to the substrate. Ca2+ and Mg2+ ions enhance the protease activity, whereas Cu2+ and Hg2+ ions were found to be inhibitory. The pivotal role of aspartic protease in initiating hemoglobin degradation in P. vivax malaria parasite is also demonstrated.

Animals↗

Electroconvulsive therapy after repair of cerebral aneurysm.

There are cases in the literature describing the use of electroconvulsive therapy (ECT) in patients with intracerebral aneurysms. We report the clinical course of a 60-year-old woman with a history of a recently ruptured intracerebral aneurysm and placement of a ventriculoperitoneal shunt that was successfully treated with ECT for major depressive disorder. Beta blockade with esmolol 20-50 mg was used to maintain her blood pressure, which ranged from 100/63 to 170/94 mm Hg. Propofol was used for anesthesia. There were no vascular complications secondary to the ECT procedure.

Aneurysm, Ruptured↗

AML1-ETO decreases ETO-2 (MTG16) interactions with nuclear receptor corepressor, an effect that impairs granulocyte differentiation.

The t(8;21) chromosome abnormality in acute myeloid leukemia targets the AML1 and ETO genes to produce the leukemia fusion protein AML1-ETO. Another member of the ETO family, ETO-2/MTG16, is highly expressed in murine and human hematopoietic cells, bears >75% homology to ETO, and like ETO, contains a conserved MYND domain that interacts with the nuclear receptor corepressor (N-CoR). AML1-ETO prevents granulocyte but not macrophage differentiation of murine 32Dcl3 granulocyte/macrophage progenitors. One possible mechanism is recruitment of N-CoR to aberrantly repress AML1 target genes. We wished to examine another mechanism by which AML1-ETO might impair granulocyte differentiation. We demonstrate that AML1-ETO decreases interactions between ETO-2 and N-CoR. Furthermore, overexpression of ETO-2 relieves AML1-ETO-induced granulocyte differentiation arrest. This suggests that decreased interactions between ETO-2 and N-CoR may contribute to granulocyte differentiation impairment. The MYND domain coimmunoprecipitates with N-CoR and inhibits interactions between ETO-2 and N-CoR, presumably by occupying the ETO-2 binding site on N-CoR. This inhibition of ETO-2 interactions with N-CoR is specific because the MYND domain does not inhibit retinoic acid receptor interactions with N-CoR. To examine the effect of decreasing interactions between ETO-2 and N-CoR in hematopoietic cells, without effects of AML1-ETO such as direct repression of AML1 target genes, the MYND domain was expressed in 32Dcl3 and human CD34+ cells. The MYND domain prevented granulocyte but not macrophage differentiation of both 32Dcl3 and human CD34+ cells, recapitulating this effect of AML1-ETO. In conclusion, decreasing interactions between ETO-2 and N-CoR, an effect of AML1-ETO, inhibits granulocyte differentiation.

Animals↗

Radiation-induced enhancement of antioxidant contents of soybean (Glycine max Merrill).

Soybean samples were treated with gamma-radiation doses between 0.5 and 5 kGy for achieving insect disinfestation and microbial decontamination. Nutritional quality of soybeans with respect to antioxidant isoflavone content was tested in radiation-treated and untreated samples. Changes in major isoflavones such as genistein, diadzein, glycetein, and their glycosides were monitored by high-performance liquid chromatography. Interestingly, a decrease in content of glycosidic conjugates and an increase in aglycons were noted with increasing radiation dose. Antioxidant potential measured as percent 1,1-diphenyl-2-picrylhydrazyl scavenging activity showed an increasing trend with dose, indicating that radiation processing as a method of food preservation has a positive nutritional implication.

Antioxidants↗