Type II taste cells participate in mucosal immune surveillance.

The oral microbiome is second only to its intestinal counterpart in diversity and abundance, but its effects on taste cells remains largely unexplored. Using single-cell RNASeq, we found that mouse taste cells, in particular, sweet and umami receptor cells that express taste 1 receptor member 3 (Tas1r3), have a gene expression signature reminiscent of Microfold (M) cells, a central player in immune surveillance in the mucosa-associated lymphoid tissue (MALT) such as those in the Peyer's patch and tonsils. Administration of tumor necrosis factor ligand superfamily member 11 (TNFSF11; also known as RANKL), a growth factor required for differentiation of M cells, dramatically increased M cell proliferation and marker gene expression in the taste papillae and in cultured taste organoids from wild-type (WT) mice. Taste papillae and organoids from knockout mice lacking Spib (SpibKO), a RANKL-regulated transcription factor required for M cell development and regeneration on the other hand, failed to respond to RANKL. Taste papillae from SpibKO mice also showed reduced expression of NF-κB signaling pathway components and proinflammatory cytokines and attracted fewer immune cells. However, lipopolysaccharide-induced expression of cytokines was strongly up-regulated in SpibKO mice compared to their WT counterparts. Like M cells, taste cells from WT but not SpibKO mice readily took up fluorescently labeled microbeads, a proxy for microbial transcytosis. The proportion of taste cell subtypes are unaltered in SpibKO mice; however, they displayed increased attraction to sweet and umami taste stimuli. We propose that taste cells are involved in immune surveillance and may tune their taste responses to microbial signaling and infection.

Value Added Conversion of Ethanol on Morphologically Controlled and Defect-Engineered Titanium Dioxide Nanorods.

Developing an environmentally benign and highly effective strategy for the value-added conversion of biomass platform molecules such as ethanol has emerged as a significant challenge and opportunity. This challenge stems from the need to harness renewable solar energy and conduct thermodynamically unfavorable reactions at room temperature. To tackle this challenge, one-dimensional titanium dioxide photocatalysts have been designed and fabricated to achieve a remarkable photocatalytic selectivity of almost 100 % for transforming ethanol into value-added 1,1-diethoxyethane, contrasting the primary production of acetaldehyde in titanium dioxide nanoparticles. By incorporating a Pt co-catalyst and infusing oxygen vacancies into the one-dimensional catalyst, the ethanol transformation rate was doubled to 128.8 mmol g-1 h-1 with respect to that of its unmodified counterpart (about 66.7 mmol g-1 h-1 ). The underlying mechanism for this high conversion and selectivity resides in the narrowed bandgap of the catalyst and the prolonged lifetime of the photo-generated carriers. This is a promising strategy for the photocatalytic transformation of essential biomass platform molecules that intertwines morphological control and defect engineering.

Crypt-like patterned electrospun nanofibrous membrane and probiotics promote intestinal epithelium models close to tissues.

In vitro intestinal epithelium models have drawn great attention to investigating intestinal biology in recent years. However, the difficulty to maintain the normal physiological status of primary intestinal epithelium in vitro limits the applications. Here, we designed patterned electrospun polylactic acid (PLA) nanofibrous membranes with crypt-like topography and mimic ECM fibrous network to support crypt culture and construct in vitro intestinal epithelium models. The patterned electrospun PLA nanofibrous membranes modified with Matrigels at 0 °C showed high biocompatibility and promoted cell growth and proliferation. The constructed duodenum epithelium models and colon epithelium models on the patterned electrospun PLA nanofibrous membranes expressed the typical differentiation markers of intestinal epithelia and the gene expression levels were close to the original tissues, especially with the help of probiotics. The constructed intestinal epithelium models could be used to assess probiotic adhesion and colonization, which were verified to show significant differences with the Caco-2 cell models due to the different cell types. These findings provide new insights and a better understanding of the roles of biophysical, biochemical, and biological signals in the construction of in vitro intestinal epithelium models as well as the potential applications of these models in the study of host-gut microbes interactions. KEY POINTS: • Patterned electrospun scaffold has crypt-like topography and ECM nanofibrous network. • Matrigels at 0°C modify scaffolds more effectively than at 37°C. • Synergy of biomimic scaffold and probiotics makes in vitro model close to tissue.

Correlation between copper speciation and transport pathway in Caco-2 cells.

BACKGROUND: Previous studies have demonstrated that, in contrast to the properties of food-derived copper, water-derived copper exerts neurotoxic effects and exhibits different speciation during digestion. The cellular uptake efficiencies of different speciation of copper are distinct. However, it is unclear whether these different speciation share the same transport pathway in intestinal epithelial cells. In the present study, the intracellular accumulation of copper derived from copper ion and copper complex solutions was investigated in Caco-2 cells. RESULTS: The cellular accumulation of copper derived from copper ions was higher than that of copper derived from the copper complex. Treatment with carboplatin and Ag+ , which are copper transporter receptor 1 (Ctr1, LC31A1) inhibitors, did not inhibit copper accumulation in Caco-2 cells, but inhibited copper accumulation in HepG2 cells. Zinc ion significantly decreased the intracellular copper content from 114 ± 7 µg g-1 protein to 88 ± 4 µg g-1 protein in the copper ion-treated Caco-2 cells, but not in the copper complex-treated Caco-2 cells (84.6 ± 14 µg g-1 protein versus 87.7 ± 20 µg g-1 protein, P > 0.05). Additionally, copper accumulation in Caco-2 and HepG2 cells significantly differed depending on different solvents (Hanks' balanced salt solution and NaNO3 , P < 0.05). CONCLUSION: These results indicate that the intracellular accumulation of copper derived from copper ion and copper complex is mediated by distinct copper transport pathways. Copper speciation may be an important factor that affects copper absorption and toxicity. © 2022 Society of Chemical Industry.

Chlorogenic acid improves intestinal morphology by enhancing intestinal stem-cell activity.

BACKGROUND: Chlorogenic acid (CGA), as one of the most abundant naturally occurring phenolic acids, has been documented to be beneficial for intestinal health. However, the underlying mechanism is still not fully understood. The adult intestinal stem cell is the critical driver of epithelial homeostasis and regeneration. RESULTS: This study hypothesized that CGA exerted intestinal health effects by modulating intestinal stem-cell functions. Lgr5-EGFP mice were treated for 14 days, and intestinal organoids derived from these mice were treated for 3 days, using CGA solution. In comparison with the control group, CGA treatment increased intestinal villous height and crypt depth in mice and augmented the area expansion and the number of budding intestinal organoids. Quantitative polymerase chain reaction (qPCR) analysis revealed that CGA treatment significantly increased the expression of genes coding intestinal stem-cell markers in intestinal tissue and organoids, and upregulated the expression of genes coding secretory cell lineages and enterocytes, although not statistically significantly. Fluorescence-activated cell-sorting analysis further confirmed that CGA augmented the number of stem cells. 5-Ethynyl-2'-deoxyuridine (EdU) incorporation and Ki67 immunostaining results also demonstrated that CGA treatment enhanced intestinal stem-cell proliferation. CONCLUSION: Altogether, our findings indicate that CGA could activate intestinal stem-cell and epithelial regeneration, which could contribute to the improvement of intestinal morphology or organoid growth of mice. This highlights a promising mechanism for CGA as an excellent candidate for the formulation of dietary supplements and functional foods for intestinal protection. © 2023 Society of Chemical Industry.

Nkx2-2 expressing taste cells in endoderm-derived taste papillae are committed to the type III lineage.

In mammals, multiple cell-signaling pathways and transcription factors regulate development of the embryonic taste system and turnover of taste cells in the adult stage. Using single-cell RNA-Seq of mouse taste cells, we found that the homeobox-containing transcription factor Nkx2-2, a target of the Sonic Hedgehog pathway and a key regulator of the development and regeneration of multiple cell types in the body, is highly expressed in type III taste cells but not in type II or taste stem cells. Using in situ hybridization and immunostaining, we confirmed that Nkx2-2 is expressed specifically in type III taste cells in the endoderm-derived circumvallate and foliate taste papillae but not in the ectoderm-derived fungiform papillae. Lineage tracing revealed that Nkx2-2-expressing cells differentiate into type III, but not type II or type I cells in circumvallate and foliate papillae. Neonatal Nkx2-2-knockout mice did not express key type III taste cell marker genes, while the expression of type II and type I taste cell marker genes were unaffected in these mice. Our findings indicate that Nkx2-2-expressing cells are committed to the type III lineage and that Nkx2-2 may be critical for the development of type III taste cells in the posterior tongue, thus illustrating a key difference in the mechanism of type III cell lineage specification between ectoderm- and endoderm-derived taste fields.

Sweet taste perception in mice is blunted by PTBP1-regulated skipping of Tas1r2 exon 4.

Taste perception, initiated by activation of taste receptors in taste bud cells, is crucial for regulating nutrient intake. Genetic polymorphisms in taste receptor genes cannot fully explain the wide individual variations of taste sensitivity. Alternative splicing (AS) is a ubiquitous posttranscriptional mode of gene regulation that enriches the functional diversity of proteins. Here, we report the identification of a novel splicing variant of sweet taste receptor gene Tas1r2 (Tas1r2_∆e4) in mouse taste buds and the mechanism by which it diminishes sweet taste responses in vitro and in vivo. Skipping of Tas1r2 exon 4 in Tas1r2_∆e4 led to loss of amino acids in the extracellular Venus flytrap domain, and the truncated isoform reduced the response of sweet taste receptors (STRs) to all sweet compounds tested by generating nonfunctional T1R2/T1R3 STR heterodimers. The splicing factor PTBP1 (polypyrimidine tract-binding protein 1) promoted Tas1r2_∆e4 generation through binding to a polypyrimidine-rich splicing silencer in Tas1r2 exon 4, thus decreasing STR function and sweet taste perception in mice. Taken together, these data reveal the existence of a regulated AS event in Tas1r2 expression and its effect on sweet taste perception, providing a novel mechanism for modulating taste sensitivity at the posttranscriptional level.

Histone deacetylase 6 promotes skin wound healing by regulating fibroblast migration and differentiation in aged mice.

Skin wound healing tends to slow down with aging, which is detrimental to both minor wound recovery in daily life and the recovery after surgery. The aim of current study was to explore the effect of histone deacetylase 6 (HDAC6) on wound healing during aging. Cultured human dermal fibroblasts (HDFs) and mouse full-thickness skin wound model were used to explore the functional changes of replicative senescent dermal fibroblasts and the effect of aging on skin wound healing. Scratch wound healing assay revealed significantly decreased migration speed of senescent HDFs, and BrdU incorporation assay indicated their considerably retardant proliferation. The protein expression levels of collagen and HDAC6 were significantly decreased in both senescent HDFs and skin tissues from aged mice. HDAC6 activity inhibition with highly selective inhibitor tubastatin A (TsA) or HDAC6 knockdown with siRNA decreased the migration speed of HDFs and considerably suppressed fibroblast differentiation induced by transforming growth factor-ß1 (TGF-ß1), which suggests the involvement of HDAC6 in regulating fundamental physiological activities of dermal fibroblasts. In vivo full-thickness skin wound healing was significantly delayed in young HDAC6 knockout mice when compared with young wild type mice. In addition, the wound healing was significantly slower in aged wild type mice than that in young wild type mice, and became even worse in aged HDAC6 knockout aged mice. Compared to the aged wild type mice, aged HDAC6 knockout mice exhibited delayed angiogenesis, reduced collagen synthesis, and decreased collagen deposition in skin wounds. Together, these results suggest that delayed skin wound healing in aged mice is associated with impaired fibroblast function. Adequate expression and activity of HDAC6 are required for fibroblasts migration and differentiation.

Effects of sunset yellow on proliferation and differentiation of intestinal epithelial cells in murine intestinal organoids.

Sunset yellow (SY), an azo dye, is commonly used in the food industry. The scientific literature contains little information regarding the effects of SY on small intestinal epithelial cells (IECs). In this study, a small intestinal organoid model was used in in vitro toxicological studies of SY, and intestinal inflammatory responses in vivo to SY were investigated with the dextran sulfate sodium (DSS)-induced intestinal inflammation model in C57BL/6 mice. The intestinal organoids were cultured with 2 µg/ml SY for two generations, the growth rates were analyzed, and the expressions of cell lineages were assayed. For inflammatory responses, mice were fed with a diet containing 40 mg/kg diet SY and treated with 2.5% DSS for 7 days. The results showed that SY inhibited the growth of the organoids by inhibiting the proliferation and disturbing the differentiation of IECs. Furthermore, endoplasmic reticulum (ER) stress and oxidative stress levels were elevated in SY-treated organoids. In DSS-treated mice, the disease activity index and expression levels of interleukin-1ß and tumor necrosis factor-α were enhanced in the SY group, concluding that SY exacerbated DSS-induced intestinal inflammation. Taken together, these findings revealed that SY could disturb the homeostasis of the small intestinal epithelium by generating high levels of ER stress and oxidative stress, with long-term continuous consumption of SY potentially increasing the risk of intestinal inflammation.

Design, synthesis, and biological evaluation of novel triazoloquinazolinone derivatives as SHP2 protein inhibitors.

A novel series of triazoloquinazolinone derivatives were designed, synthesised, and evaluated for their in vitro biological activities against the SHP2 protein. Moreover, some compounds were evaluated against A375 cells. The results revealed that target compounds possessed moderate to excellent inhibitory activity against SHP2 protein, whereas compounds 12f, 12l, 12j, 17e, and 17f have strong antiproliferative activity on A375 cells. The compound 12l showed remarkable cytotoxicity against A375 cells and a strong inhibitory effect against SHP2 protein when compared with SHP244. The structure-activity relationships (SARs) indicated that electron-donating groups (EDGs) on phenyl rings are beneficial for improving the antitumor activity; compounds with a hydroxyl substituent at the 2-position of phenyl ring exhibited superior activities than compounds with a substituent at the 4-position. In addition, compound 12l displayed improved physicochemical properties as well as metabolic stability compared to SHP244. Our efforts identified 12l as a promising SHP2 protein inhibitor, warranting its further investigation.

Gli3 is a negative regulator of Tas1r3-expressing taste cells.

Mouse taste receptor cells survive from 3-24 days, necessitating their regeneration throughout adulthood. In anterior tongue, sonic hedgehog (SHH), released by a subpopulation of basal taste cells, regulates transcription factors Gli2 and Gli3 in stem cells to control taste cell regeneration. Using single-cell RNA-Seq we found that Gli3 is highly expressed in Tas1r3-expressing taste receptor cells and Lgr5+ taste stem cells in posterior tongue. By PCR and immunohistochemistry we found that Gli3 was expressed in taste buds in all taste fields. Conditional knockout mice lacking Gli3 in the posterior tongue (Gli3CKO) had larger taste buds containing more taste cells than did control wild-type (Gli3WT) mice. In comparison to wild-type mice, Gli3CKO mice had more Lgr5+ and Tas1r3+ cells, but fewer type III cells. Similar changes were observed ex vivo in Gli3CKO taste organoids cultured from Lgr5+ taste stem cells. Further, the expression of several taste marker and Gli3 target genes was altered in Gli3CKO mice and/or organoids. Mirroring these changes, Gli3CKO mice had increased lick responses to sweet and umami stimuli, decreased lick responses to bitter and sour taste stimuli, and increased glossopharyngeal taste nerve responses to sweet and bitter compounds. Our results indicate that Gli3 is a suppressor of stem cell proliferation that affects the number and function of mature taste cells, especially Tas1r3+ cells, in adult posterior tongue. Our findings shed light on the role of the Shh pathway in adult taste cell regeneration and may help devise strategies for treating taste distortions from chemotherapy and aging.

Chronic administration of caffeine alters acesulfame-K intake and features of fungiform taste buds in mice.

The objective of this study was to investigate the effects of chronic administration of caffeine on the anatomical characteristics of taste buds, the expression level of taste receptor protein in mice, and preference for a palatable solution. We found that following a 21-day administration of caffeine, mice showed increased behavioural responses to sweet stimuli (acesulfame-K solution). Mirroring this behavioural change, chronic caffeine treatment evidently decreased the maximal cross-sectional area and height of the longitudinal axis of fungiform taste buds, the number of taste cells per fungiform taste bud, and the expression of G protein α-gustducin, while the expression of the sweet taste receptors T1R2 and T1R3 was reversed. Our findings demonstrate that chronic administration of caffeine has an impact on taste sensitivity and changes in taste bud features, which may contribute to the alteration of taste behaviour.

An optimized organic acid human sensory sourness analysis method.

BACKGROUND: Sour taste perception builds on both chemical and physiological foundations, and plays an important role in food flavor, including that of fruit, beer, wine, and other beverages. A uniform sourness standard and sourness conversion method for researchers and food enterprises is necessary to obtain uniform conclusions. RESULTS: This study established an optimized organic acid sensory sourness analysis and sourness conversion method. It is based on sour sensory difference strength curves, which consist of an absolute threshold value and sensory difference threshold values. Defining the absolute threshold value of citric acid sourness as 1, sourness could be calculated according to the curve. With a logarithmic curve form, the acid sourness indexes (AI) were calculated as 1, 0.74, 0.77, 1.31, and 1.21 for citric, malic, fumaric, lactic, and tartaric acid samples, respectively. Consequently, each acid's sourness and concentration could be obtained and converted. Single acid and mixed acid sourness comparison evaluation's result implied that the novel method was more accurate (91.7-100%) than the hydrogen ion concentration method. CONCLUSION: The novel sourness determination and conversion equation would provide more accurate sourness standard and calculation method in food sensory areas. © 2021 Society of Chemical Industry.

A Broad-Spectrum Sweet Taste Sensor Based on Ni(OH)2/Ni Electrode.

A broad-spectrum sweet taste sensor based on Ni(OH)2/Ni electrode was fabricated by the cyclic voltammetry technique. This sensor can be directly used to detect natural sweet substances in 0.1 M NaOH solution by chronoamperometry method. The current value measured by the sensor shows a linear relationship with the concentration of glucose, sucrose, fructose, maltose, lactose, xylitol, sorbitol, and erythritol (R² = 0.998, 0.983, 0.999, 0.989, 0.985, 0.990, 0.991, and 0.985, respectively). Moreover, the characteristic value of this sensor is well correlated with the concentration and relative sweetness of eight sweet substances. The good correlation between the characteristic value of six fruit samples measured by the sensor and human sensory sweetness measured by sensory evaluation (correlation coefficient = 0.95) indicates that it can reflect the sweetness of fruits containing several sweet substances. In addition, the sensor also exhibits good long-term stability over 40 days (signal ratio fluctuation ranges from 91.5% to 116.2%). Thus, this broad-spectrum sensor is promising for sweet taste sensory application.

8-Anilino-1-naphthalenesulfonate/Layered Double Hydroxide Ultrathin Films: Small Anion Assembly and Its Potential Application as a Fluorescent Biosensor.

The fluorescent dye 8-anilino-1-naphthalenesulfonate (ANS) is a widely used fluorescent probe molecule for biochemistry analysis. This paper reported the fabrication of ANS/layered double hydroxide nanosheets (ANS/LDH)n ultrathin films (UTFs) via the layer-by-layer small anion assembly technique based on electrostatic interaction and two possible weak interactions: hydrogen-bond and induced electrostatic interactions between ANS and positive-charged LDH nanosheets. The obtained UTFs show a long-range-ordered periodic layered stacking structure and weak fluorescence in dry air or water, but it split into three narrow strong peaks in a weak polarity environment induced by the two-dimensional (2D) confinement effect of the LDH laminate; the fluorescence intensity increases with decreasing the solvent polarity, concomitant with the blue shift of the emission peaks, which show good sensoring reversibility. Meanwhile, the UTFs exhibit selective fluorescence enhancement to the bovine serum albumin (BSA)-like protein biomolecules, and the rate of fluorescence enhancement with the protein concentration is significantly different with the different protein aggregate states. The (ANS/LDH)n UTF has the potential to be a novel type of biological flourescence sensor material.

Diminishing Self-Discharge of High-Loading Li-S Batteries with Oxygen-Rich Biomass Carbon Interlayers.

Lithium-sulfur batteries (Li-S) have possessed gratifying development in the past decade due to their high theoretical energy density. However, the severe polysulfide shuttling provokes undesirable self-discharge effect, leading to low energy efficiency in Li-S batteries. Herein, an interlayer composed of oxygen-rich carbon nanosheets (OCN) derived from bagasse is elaborated to suppress the shuttle effect and reduce the resultant self-discharge effect. The OCN interlayer is able to physically block the shuttling behavior of polysulfides and its oxygen-rich functional groups can strongly interact with polysulfides via O-S bonds to chemically immobilize mobile polysulfides. The self-discharge test for seven days further shows that the self-discahrge rate is diminished by impressive 93 %. As a result, Li-S batteries with the OCN interlayer achieve an ultrahigh discharge specific capacity of 710 mAh g-1 at a high mass loading of 7.18 mg. The work provides a facile method for designing functional interlayers and opens a new avenue for realizing Li-S batteries with high energy efficiency.

The novel lactoferrin and DHA-codelivered liposomes with different membrane structures: Fabrication, in vitro infant digestion, and suckling pig intestinal organoid absorption.

Inspired by membrane structure of breast milk and infant formula fat globules, four liposomes with different particle size (large and small) and compositions (Single phospholipids contained phosphatidylcholine, complex phospholipids contained phosphatidylcholine, phosphatidylethanolamine and sphingomyelin) were fabricated to deliver lactoferrin and DHA. In vitro infant semi-dynamic digestive behavior and absorption in intestinal organoids of liposomes were investigated. Liposomal structures were negligible changed during semi-dynamic gastric digestion while damaged in intestine. Liposomal degradation rate was primarily influenced by particle size, and complex phospholipids accelerated DHA hydrolysis. The release rate of DHA (91.7 ± 1.3 %) in small-sized liposomes (0.181 ± 0.001 µm) was higher than free DHA (unencapsulated, 64.6 ± 3.4 %). Complex phospholipids liposomal digesta exhibited higher transport efficiency (3.4-fold for fatty acids and 2.0-fold for amino acids) and better organoid growth than digesta of bare nutrients. This study provided new insights into membrane structure-functionality relationship of liposomes and may aid in the development of novel infant nutrient carriers.

A strategy for mechanically integrating robust hydrogel-tissue hybrid to promote the anti-calcification and endothelialization of bioprosthetic heart valve.

Bioprosthetic heart valve (BHV) replacement has been the predominant treatment for severe heart valve diseases over decades. Most clinically available BHVs are crosslinked by glutaraldehyde (GLUT), while the high toxicity of residual GLUT could initiate calcification, severe thrombosis, and delayed endothelialization. Here, we construed a mechanically integrating robust hydrogel-tissue hybrid to improve the performance of BHVs. In particular, recombinant humanized collagen type III (rhCOLIII), which was precisely customized with anti-coagulant and pro-endothelialization bioactivity, was first incorporated into the polyvinyl alcohol (PVA)-based hydrogel via hydrogen bond interactions. Then, tannic acid was introduced to enhance the mechanical performance of PVA-based hydrogel and interfacial bonding between the hydrogel layer and bio-derived tissue due to the strong affinity for a wide range of substrates. In vitro and in vivo experimental results confirmed that the GLUT-crosslinked BHVs modified by the robust PVA-based hydrogel embedded rhCOLIII and TA possessed long-term anti-coagulant, accelerated endothelialization, mild inflammatory response and anti-calcification properties. Therefore, our mechanically integrating robust hydrogel-tissue hybrid strategy showed the potential to enhance the service function and prolong the service life of the BHVs after implantation.

A drug-free cardiovascular stent functionalized with tailored collagen supports in-situ healing of vascular tissues.

Drug-eluting stent implantation suppresses the excessive proliferation of smooth muscle cells to reduce in-stent restenosis. However, the efficacy of drug-eluting stents remains limited due to delayed reendothelialization, impaired intimal remodeling, and potentially increased late restenosis. Here, we show that a drug-free coating formulation functionalized with tailored recombinant humanized type III collagen exerts one-produces-multi effects in response to injured tissue following stent implantation. We demonstrate that the one-produces-multi coating possesses anticoagulation, anti-inflammatory, and intimal hyperplasia suppression properties. We perform transcriptome analysis to indicate that the drug-free coating favors the endothelialization process and induces the conversion of smooth muscle cells to a contractile phenotype. We find that compared to drug-eluting stents, our drug-free stent reduces in-stent restenosis in rabbit and porcine models and improves vascular neointimal healing in a rabbit model. Collectively, the one-produces-multi drug-free system represents a promising strategy for the next-generation of stents.

Effects of early intraoral acesulfame-K stimulation to mice on the adult's sweet preference and the expression of α-gustducin in fungiform papilla.

Exposure to artificial sweetener acesulfame-K (AK) at early development stages may influence the adult sweet preference and the periphery gustatory system. We observed that the intraoral AK stimulation to mice from postnatal day 4 (P4) to weaning decreased the preference thresholds for AK and sucrose solutions in adulthood, with the preference pattern unchanged. The preference scores were increased in the exposure group significantly when compared with the control group at a range of concentrations for AK or sucrose solution. Meanwhile, more α-Gustducin-labeled fungiform taste buds and cells in a single taste bud were induced from week 7 by the early intraoral AK stimulation. However, the growth in the number of α-Gustducin-positive taste bud or positive cell number per taste bud occurred only in the anterior region, the rostral 1-mm part, but not in the intermediate region, the caudal 4-mm part, of the anterior two-third of the tongue containing fungiform papillae. This work extends our previous observations and provides new information about the developmental and regional expression pattern of α-Gustducin in mouse fungiform taste bud under early AK-stimulated conditions.