CIELAB Color Space as a Field for Tracking Color-Changing Chemical Reactions of Polymeric pH Indicators.

Two types of cross-linked polymeric dye films that exhibit reversible color changes in response to pH were prepared. Upon protonation, films with a nitrophenol dye moiety (NP) changed from yellow to colorless, whereas films with an azobenzene dye moiety (AB) changed from yellow to reddish-purple. The colored light transmitted through these films from a D65 white-light source was measured colorimetrically, and the transition of the chromaticity point in CIELAB upon protonation of the polymeric dyes was observed. At low dye concentrations ([NP] < 4.7 × 10-5 and [AB] < 2.7 × 10-5 mol L-1), linear loci of the chromaticity points were displayed for both polymeric dye films, indicating perceptual linearity. This linearity is caused by the avoidance of nonlinear perceptual phenomena such as the Abney effect and the Bezold-Brücke phenomenon. The perceptual linearity of space is a necessary condition for achieving stoichiometric linearity, which requires an additional linear relationship between the dye concentration and each component, a*, b*, and L*. A geometric equation that included the AB concentration and the protonation ratio of AB was used to successfully depict, on the basis of stoichiometric linearity, the protonation of the polymeric AB films three-dimensionally in the restricted area of CIELAB.

Long-term Major Adverse Cardiac Event Prediction by Computed Tomography-derived Plaque Measures and Clinical Parameters Using Machine Learning.

Objectives The present study evaluated the usefulness of machine learning (ML) models with the coronary artery calcification score (CACS) and clinical parameters for predicting major adverse cardiac events (MACEs). Methods The Nationwide Gender-specific Atherosclerosis Determinants Estimation and Ischemic Cardiovascular Disease Prospective Cohort study (NADESICO) of 1,187 patients with suspected coronary artery disease (CAD) 50-74 years old was used to build a MACE prediction model. The ML random forest (RF) model was compared with a logistic regression analysis. The performance of the ML model was evaluated using the area under the curve (AUC) with the 95% confidence interval (CI). Results Among 1,178 patients from the NADESICO dataset, MACEs occurred in 103 (8.7%) patients during a median follow-up of 4.4 years. The AUC of the RF model for MACE prediction was 0.781 (95% CI: 0.670-0.870), which was significantly higher than that of the conventional logistic regression model [AUC, 0.750 (95% CI, 0.651-0.839)]. The important features in the RF model were coronary artery stenosis (CAS) at any site, CAS in the left anterior descending branch, HbA1c level, CAS in the right coronary artery, and sex. In the external validation cohort, the model accuracy of ensemble ML-RF models that were trained on and tuned using the NADESICO dataset was not similar [AUC: 0.635 (95% CI: 0.599-0.672)]. Conclusion The ML-RF model improved the long-term prediction of MACEs compared to the logistic regression model. However, the selected variables in the internal dataset were not highly predictive of the external dataset. Further investigations are required to validate the usefulness of this model.

PGC-based cryobanking, regeneration through germline chimera mating, and CRISPR/Cas9-mediated TYRP1 modification in indigenous Chinese chickens.

Primordial germ cells (PGCs) are vital for producing sperm and eggs and are crucial for conserving chicken germplasm and creating genetically modified chickens. However, efforts to use PGCs for preserving native chicken germplasm and genetic modification via CRISPR/Cas9 are limited. Here we show that we established 289 PGC lines from eight Chinese chicken populations with an 81.6% success rate. We regenerated Piao chickens by repropagating cryopreserved PGCs and transplanting them into recipient chickens, achieving a 12.7% efficiency rate. These regenerated chickens carried mitochondrial DNA from female donor PGC and the rumplessness mutation from both male and female donors. Additionally, we created the TYRP1 (tyrosinase-related protein 1) knockout (KO) PGC lines via CRISPR/Cas9. Transplanting KO cells into male recipients and mating them with wild-type hens produced four TYRP1 KO chickens with brown plumage due to reduced eumelanin production. Our work demonstrates efficient PGC culture, cryopreservation, regeneration, and gene editing in chickens.

Infusion Reactions in HER2-Positive Gastric Cancer: Switching from Trastuzumab to Its Biosimilar.

This study presents a safety analysis of infusion reactions (IRs) in gastric cancer patients who switched from reference trastuzumab to its biosimilar, trastuzumab-NK, at the Saitama Cancer Center in Japan from April 2018 to March 2022. IRs were identified if patients developed symptoms such as fever, chills, infusion-related reactions, hypersensitivity, rash, pruritus, urticaria, systemic disorders, or immune system disorders on the day of administration or the following day. The incidence of IRs was 14% in the reference trastuzumab group, 33% in the trastuzumab-NK group, and 33% in the switching group. There was no significant difference in IR incidence between the reference trastuzumab and trastuzumab-NK groups (p = 0.235). Among the switching group, only one of the three patients who experienced an IR had a reaction associated with the switch. These findings suggest that the frequency of IRs in the switching group gastric cancer is comparable to the other groups, indicating that switching is a viable treatment option with appropriate management. Additionally, 37 of the 45 patients in the study were male, provides new safety information on switching in gastric cancer for male patients that has not been previously reported.

A systematic exploration of unexploited genes for oxidative stress in Parkinson's disease.

Human disease-associated gene data are accessible through databases, including the Open Targets Platform, DisGeNET, miRTex, RNADisease, and PubChem. However, missing data entries in such databases are anticipated because of curational errors, biases, and text-mining failures. Additionally, the extensive research on human diseases has led to challenges in registering comprehensive data. The lack of essential data in databases hinders knowledge sharing and should be addressed. Therefore, we propose an analysis pipeline to explore missing entries of unexploited genes in the human disease-associated gene databases. Using this pipeline for genes in Parkinson's disease with oxidative stress revealed two unexploited genes: nuclear protein 1 (NUPR1) and ubiquitin-like with PHD and ring finger domains 2 (UHRF2). This methodology enhances the identification of underrepresented disease-associated genes, facilitating easier access to potential human disease-related functional genes. This study aims to identify unexploited genes for further research and does not include independent experimental validation.

The Predictive Risk Factor of Postoperative Recurrence Following Altemeier's and Delorme's Procedures for Full-thickness Rectal Prolapse: An Analysis of 127 Japanese Patients in a Single Institution.

Objectives: We aimed to identify risk factors for postoperative recurrence (PR) after Altemeier's and Delorme's procedures for full-thickness rectal prolapse (FTRP). Methods: We enrolled 127 patients who underwent Altemeier's and Delorme's procedures for FTRP between April 2008 and September 2021. We divided the 127 patients into recurrence and non-recurrence groups and conducted univariate and multivariate analyses. We used six independent variables: age, body mass index (BMI), history of surgical repair for FTRP, coexistence of prolapse in other organs, poor fixation of the rectum on defecography before surgery, length of the prolapsed rectum, and type of surgical procedure (Altemeier's or Delorme's procedures). Results: PR developed in 51 (40.1%) patients during a mean follow-up period of 453 (range, 9-3616) days. Comparing the recurrence group (n=51) with the non-recurrence group (n=76), significant difference was observed regarding the coexistence of prolapse in other organs (p=0.017) in the univariate analysis. In the multivariate analysis, significant differences were observed in BMI (OR 1.18, 95% CI 1.030-1.350, p=0.020), coexistence of prolapse in other organs (OR 3.38, 95% CI 1.200-9.500, p=0.021), length of the prolapsed rectum (OR 1.030, 95% CI 1.010-1.060, p=0.015), poor fixity of the rectum on defecography (OR 0.332, 95% CI 0.129-0.852, p=0.022), and surgical procedures (OR 0.192, 95% CI 0.064-0.573, p=0.003). Conclusions: The study suggested that increasing BMI, coexistence of prolapse in other organs, length of the prolapsed rectum, poor fixation of the rectum on defecography before surgery, and types of surgical procedure might be risk factors of PR after perineal surgery for FTRP.

Vasa vasorum enhancement on optical coherence tomography in Kawasaki disease.

BACKGROUND: Patients with Kawasaki disease (KD) prone to develop coronary artery aneurysm (CAA) with unknown etiology. We aimed to disclose the relationship between vasa vasorum (VV) and intimal thickening using optical coherence tomography (OCT) in KD. METHODS: Forty-three coronary artery branches of 21 patients with KD were examined by OCT. The coronary arteries were classified into three groups: the CAA group (n = 9) in which CAAs remained since the acute phase, the regressed group (n = 16) in which CAAs were regressed, and the no CAA group (n = 18). The number and distribution of VV, and intimal thickening in coronary arteries were evaluated on OCT. RESULTS: Intimal thickening was significantly more severe in the CAA and regressed groups than in the no CAA group (median: 481, 474, and 218 µm, p = 0.001 and p < 0.001, respectively). The number of VV in the regressed group was significantly higher than that in the CAA and no CAA groups. The numbers of adventitial VV and internal VV were positively correlated with the intimal thickness (R = 0.64, p < 0.001; R = 0.62, p < 0.001, respectively). In the no CAA group, no internal VV were observed. CONCLUSIONS: VV enhances according to intimal thickening, suggesting that VV may have some link to the healing process, such as CAA regression and intimal thickening. IMPACT: Kawasaki disease (KD) is a vasculitis syndrome developing coronary artery aneurysm, however its etiology still remains unclear. Coronary artery imaging using optical coherence tomography (OCT) can reveal coronary arterial wall pathology, however OCT studies are limited in patients with KD. Using OCT, we disclosed the closed relationship between vasa vasorum enhancement and regressed coronary arterial lesions. Vasa vasorum enhancement is involved in the pathomechanism of the convalescent phase of KD.

Pseudoirreversible inhibition elicits persistent efficacy of a sphingosine 1-phosphate receptor 1 antagonist.

Sphingosine 1-phosphate receptor 1 (S1PR1), a G protein-coupled receptor, is required for lymphocyte trafficking, and is a promising therapeutic target in inflammatory diseases. Here, we synthesize a competitive S1PR1 antagonist, KSI-6666, that effectively suppresses pathogenic inflammation. Metadynamics simulations suggest that the interaction of KSI-6666 with a methionine residue Met124 in the ligand-binding pocket of S1PR1 may inhibit the dissociation of KSI-6666 from S1PR1. Consistently, in vitro functional and mutational analyses reveal that KSI-6666 causes pseudoirreversible inhibition of S1PR1, dependent on the Met124 of the protein and substituents on the distal benzene ring of KSI-6666. Moreover, in vivo study suggests that this pseudoirreversible inhibition is responsible for the persistent activity of KSI-6666.

A Rare Case of Ileocecal Lymph Node Recurrence After Surgery in Siewert's Classification Type I Esophagogastric Junction Adenocarcinoma.

BACKGROUND Although recurrence after surgery for esophagogastric junction (EGJ) adenocarcinoma frequently develops in the mediastinal and para-aortic lymph nodes (LN), distant LN recurrence in the mesocolon is rare. We report a rare case of ileocecal LN metastasis in the ascending mesocolon after radical surgery for an EGJ adenocarcinoma. CASE REPORT We performed subtotal esophagectomy with mediastinal and para-gastric LN dissection in a patient with an advanced EGJ adenocarcinoma. Clinicopathologically, the patient was diagnosed with type I EGJ adenocarcinoma based on Siewert's classification (pathological T3N1M0). One year after surgery, computed tomography showed enlarged lymph nodes around the ileocolic artery, and further examination was performed. Although positron emission tomography-computed tomography showed that the lesion had moderate uptake of fluorodeoxyglucose, we did not find the reason for the enlarged lymph nodes. Finally, laparoscopic ileocecal resection was performed for diagnostic and therapeutic purposes. Clinicopathological tests revealed that the specimen was a moderately differentiated adenocarcinoma, which was strongly suspected to be a metastasis of the EGJ adenocarcinoma. CONCLUSIONS We encountered a rare case of EGJ adenocarcinoma that spread to the ileocecal LN in the ascending mesocolon. To the best of our knowledge, this is the first such report in the literature to date. Laparoscopic ileocecal resection for metastasis to the ascending mesocolon seems reasonable as a local control.

Identifying key morphometrics to post-storm beach recovery through explainable AI.

In the context of ongoing discussions about climate change, the focus on beach resilience has gained significant attention in contemporary studies. However, a comprehensive understanding of beach resilience, particularly in the short term, remains limited. This study utilizes a dataset of 104 storm events in Hasaki beach, located on the East coast of Japan, to investigate the 10-day beach recovery. The study considers four types of distinct beach profile patterns based on sandbar formations. Employing XGBoost and the SHAP explanation method, the influence of morphometric indicators on beach resilience were explored. Resilient beach profiles are anticipated to exhibit rapid recovery following erosional storm events. The results reveal that morphometrics play a crucial role in determining the short-term, 10-day, recovery of beaches, with specific morphometric features demonstrating pronounced effects based on profile patterns. The study contributes to the current knowledge of post-storm beach recovery and provides insights that could inform discussions on beach resilience.

Pathological Alterations of Coronary Arteries Late After Kawasaki Disease: An Optical Coherence Tomography Study.

Background: The long-term impact of Kawasaki disease on coronary arteries in vivo is unclear. Objectives: The purpose of this study was to investigate coronary arteries in the late convalescent phase, we followed patients with Kawasaki disease who developed coronary artery aneurysms (CAAs). Methods: We followed 24 patients and used optical coherence tomography at a median of 16.6 years after the onset of Kawasaki disease. Results: Of 72 coronary arteries, optical coherence tomography was performed on 61 arteries: 17 with a persistent CAA, 29 with a regressed CAA, and 15 without a CAA. Between-group comparison was performed by chi-square or Fisher's exact test, and intimal thickening (17 vs 29 vs 15, all 100%, P = NA) and medial disruption (17 [100%] vs 29 [100%] vs 14 [93%], P = 0.25) were commonly observed in the investigated arteries. Advanced features of atherosclerosis were more frequently seen in arteries with persistent CAAs than in those with regressed CAAs and in those without CAAs: calcification (12 [71%] vs 5 [17%] vs 1 [7%], P < 0.001), microvessels (12 [71%] vs 10 [35%] vs 4 [27%], P = 0.020), cholesterol crystals (6 [35%] vs 2 [7%] vs 0 [0%], P = 0.009), macrophage accumulation (11 [65%] vs 4 [14%] vs 4 [27%], P = 0.002), and layered plaque (8 [47%] vs 11 [38%] vs 0 [0%], P = 0.004). Conclusions: Long after onset of Kawasaki disease, all arteries showed pathological changes. Arteries with persistent CAAs had more advanced features of atherosclerosis than those with regressed CAAs and those without CAAs.

Preoperative Low Creatine Kinase as a Poor Prognostic Factor in Patients with Colorectal Cancer.

PURPOSE: This study aimed to evaluate the clinicopathological and prognostic significance of preoperative serum creatine kinase (CK) levels in colorectal cancer. METHODS: This study analyzed 1169 patients with colorectal cancer at stages 0 (n = 35), I (n = 301), II (n = 456), III (n = 339), and IV (n = 38). The CK cut-off value was 52 U/L to predict recurrence based on receiver operative characteristics curve. Clinicopathological factors were compared between the low (< 52 U/L) and high CK groups (≥ 52 U/L). The multivariate analysis evaluated relapse-free survival (RFS) and overall survival (OS) following CK status. RESULTS: The female sex, elderly age (≥ 75), deep tumor (pT4), and carcinoembryonic antigen (+) were independently associated with low CK status. The recurrent rate was significantly higher in the low CK group than in the high CK group (19.1% vs. 11.7%, p < 0.001). Elderly age, pT4, pN (+), preoperative carbohydrate antigen (CA) 19-9 (+), and low CK status were independent risk factors for RFS. Elderly age, pT4, pN (+), preoperative CA19-9 (+), and low CK status were independent risk factors for OS. CONCLUSION: Preoperative low CK status was associated with deep tumors and was a poor prognostic factor in patients with colorectal cancer.

Laser speckle flowgraphy has comparable accuracy to indocyanine green fluorescence angiography in assessing bowel blood perfusion.

PURPOSE: To investigate the accuracy of laser speckle flowgraphy (LSFG), a noninvasive method for the quantitative evaluation of blood flow using mean blur rate (MBR) as a blood flow parameter in the assessment of bowel blood perfusion compared to indocyanine green fluorescence angiography (ICG-FA). METHODS: We enrolled 46 patients who underwent left-sided colorectal surgery. LSFG and ICG-FA were applied to assess blood bowel perfusion, with MBR and luminance as parameters, respectively. In both measurement methods, the position where the parameter suddenly decreased was defined as the blood flow boundary line. Subsequently, the blood flow boundaries created after processing the blood vessels flowing into the intestinal tract were determined using LSFG and ICG-FA, and concordance between the two was examined. Blood flow boundaries were visually identified using color tone changes on a color map created based on MBR in LSFG and using differences in luminance in ICG-FA. The distances between the transection line and blood flow boundaries determined using each method were compared. RESULTS: The location of blood flow boundaries matched in 65% (30/46) of cases. Although locations differed in the remaining 35% (16/46), all were located on the anal side near the transection line, and the difference was not clinically significant. The average distances between the transection line and blood flow boundary were 2.76 (SD = 3.25) and 3.71 (SD = 4.26) mm, respectively. There was no statistically significant difference between the two groups (p = 0.38). CONCLUSION: LSFG was shown to have comparable accuracy to ICG-FA, and may be useful for evaluating bowel perfusion.

A Case of Type 1 Diabetes Mellitus with Diabetic Ketoacidosis Presenting with Transient Severe Fatty Liver and Subsequent Liver Dysfunction.

Patients often present with severe fatty liver (FL) due to insulin deficiency at the onset of diabetic ketoacidosis (DKA). On the other hand, glycogenic hepatopathy (GH) is a possible cause of liver dysfunction in patients with DKA. We herein report a case of type 1 diabetes mellitus with severe FL at the onset of DKA, who demonstrated subsequent marked liver dysfunction after achieving an improvement of FL. As liver dysfunction persisted even after the FL improved, GH was suspected to be the cause of liver dysfunction. FL and GH have different prognoses and should therefore be differentiated using imaging studies and biopsies.

Functional analysis of a first hindlimb positioning enhancer via Gdf11 expression.

During the early development of tetrapods, including humans, the embryonic body elongates caudally once the anterior-posterior axis is established. During this process, region-specific vertebral morphogenesis occurs, with the determination of limb positioning along the anterior-posterior axis. We previously reported that Gdf11 functions as an anatomical integration system that determines the positioning of hindlimbs and sacral vertebrae where Gdf11 is expressed. However, the molecular mechanisms underlying induction of Gdf11 expression remain unclear. In this study, we searched for non-coding regions near the Gdf11 locus that were conserved across species to elucidate the regulatory mechanisms of Gdf11 expression. We identified an enhancer of the Gdf11 gene in intron 1 and named it highly conserved region (HCR). In HCR knockout mice, the expression level of endogenous Gdf11 was decreased, and the position of the sacral-hindlimb unit was shifted posteriorly. We also searched for factors upstream of Gdf11 based on the predicted transcription factor binding sites within the HCR. We found that inhibition of FGF signaling increased endogenous Gdf11 expression, suggesting that FGF signaling negatively regulates Gdf11 expression. However, FGF signaling does not regulate HCR activity. Our results suggest that there are species-specific Gdf11 enhancers other than HCR and that FGF signaling regulates Gdf11 expression independent of HCR.

Accuracy of aortic valve calcification volume score for identification of significant aortic stenosis on non-electrocardiographic-gated computed tomography compared to the Agatston scoring system.

BACKGROUND: Considering the absence of reports validating the precision of the volume score and the relationship between the volume and Agatston scores, this study evaluated the accuracy of the volume score compared to the Agatston score for the quantitative measurement of aortic valve calcification (AVC) on non-electrocardiographic-gated computed tomography (CT). METHODS: We retrospectively analysed the AVC scores of 5385 patients who underwent transthoracic echocardiography between March 1, 2013 and December 26, 2019 â€‹at our institution, using non-contrast non-electrocardiographic-gated CT. The thresholds for significant aortic stenosis (AS) were computed using receiver operating characteristic curves based on the AVC scores. The area under the curve (AUC) of the Agatston and volume scores for significant AS were compared to evaluate the accuracy of the scoring method. RESULTS: All sex-specific AVC thresholds of the volume score for significant AS (moderate and high AS severity, moderate and high AS severity without discordance, discordant severe AS, and concordant severe AS) showed high sensitivity and specificity (AUC, 0.978-0.996; sensitivity, 94.2-98.4%; specificity, 90.1-100%). No significant differences in the AUC were observed between the Agatston and volume scores for significant AS in male and female patients. CONCLUSION: All volume score threshold values showed high sensitivity and specificity for identifying significant AS. The accuracy of the test for AVC thresholds of the volume score for significant AS was comparable to that of the Agatston score. Our findings raise questions about the significance of weighting calcium density in the Agatston score for assessing AS severity.

Successful retrieval of a foreign body in an infant's right pulmonary artery using the new boomerang loop-snare technique: A case report.

We report successful percutaneous retrieval of a foreign body located in an infant's right pulmonary artery using the new boomerang loop-snare technique. The case was an 18-month-old girl. A central venous catheter for chemotherapy was inserted from the right subclavian vein during treatment for myelodysplastic syndrome at another hospital. A postprocedural chest X-ray showed a foreign body in her right lung, and contrast-enhanced computed tomography confirmed the linear foreign body was located in the right pulmonary artery. The patient was transferred to our hospital to retrieve the foreign body. Under cooperation with pediatric cardiologists, a 6 Fr sheath was inserted via the right femoral vein, and a guiding catheter was advanced into the right pulmonary artery. Owing to the risk of vascular injury when using endoscopic forceps, we decided to use the loop-snare technique. We successfully crossed over the foreign body using a steerable microcatheter and a long microguidewire. The microguidewire was reinserted into the guiding catheter, and a loop was created by grasping the end of the wire using a microsnare catheter, which was inserted coaxially within the guiding catheter. By pulling the microsnare catheter, we were able to pull the foreign body into the guiding catheter and successfully retrieved it. There were no complications, such as pulmonary artery injuries or thrombi. The recovered foreign body was a piece of a guidewire. The boomerang loop-snare technique using a small-diameter system is useful for the retrieval of a foreign body in infants.

Differentiating gallbladder cancer from polyps using non-enhanced magnetic resonance imaging.

Purpose: This retrospective cohort study assessed the efficiency of non-enhanced magnetic resonance imaging (MRI) for differentiating gallbladder cancer (GBC) from gallbladder polyps (GBPs) measuring ≥ 10 mm. Material and methods: Patients diagnosed with GBCs or GBPs ≥ 10 mm and GBC ≤ T2 stage were eligible for inclusion. Two independent blinded readers assessed the continuity of the mucosal and muscular layers (CMML; present or absent) and normalised signal intensity ratio (NIR) on the apparent diffusion coefficient map (NIR-ADC), T1-weighted image (NIR-T1WI), and T2-weighted half-Fourier acquisition single-shot turbo spin-echo image. Univariate and multivariate logistic regression analyses and interobserver agreement analyses were performed to detect predictive variables differentiating GBCs from GBPs. Receiver operating characteristic (ROC) analysis was performed to evaluate diagnostic performance. A reproducibility test was performed to verify the predictive variables. Results: Multivariate analysis showed significant differences in CMML, NIR-ADC, and NIR-T1WI (p < 0.001). The positive predictive value (PPV) and specificity of the absence of CMML were approximately 100%. The CMML showed the best specificity, accuracy, and PPV in the reproducibility study. The sensitivity of CMML alone was approximately 50%, whereas it increased to approximately 70% when combined with NIR-ADC. The diagnostic performance of the combination, including sensitivity, was almost like that of tumour size. The combined tumour size and CMML assessment showed higher diagnostic performance than tumour size alone. Conclusions: The absence of CMML and NIR-ADC ≤ 1.86 helped in differentiating GBCs from GBPs. Evaluation of the absence of CMML and measurement of tumour size could better aid in determining between the two than measurement of tumour size alone.

Evaluation of the Simple Suspension Method for Oral Anticancer Drugs.

The simple suspension method (SSM) involves administering tablets or capsules using a tube after disintegration and suspension in hot water without crushing or opening the capsule. Particularly, for anticancer drugs, it is an excellent method of administration that reduces the risk of exposure during dispensing. In contrast, information on SSM for individual drugs is insufficient. Anticancer drugs present a therapeutic challenge because their information is limited. We investigated whether SSM is possible with 36 anticancer drugs. Furthermore, we examined the pH of the suspension of these drugs, for which no information on SSM is available. We found that suspension was possible for 24 of the 36 drugs. Furthermore, the pH of the suspension was measured, which provided important information when considering dissolution solutions other than hot water. Little changes in the pH were observed before or after passing through the tube. The results of this study may improve medication adherence in patients with cancer experiencing dysphagia.

Current research on mechanisms of limb bud development, and challenges for the next decade.

The developmental mechanisms of limb buds have been studied in developmental biology as an excellent model of pattern formation. Chick embryos have contributed to the discovery of new principles in developmental biology, as it is easy to observe live embryos and manipulate embryonic tissues. Herein, I outline recent findings and future issues over the next decade regarding three themes, based on my research: limb positioning, proximal-distal limb elongation and digit identity determination. First, how hindlimb position is determined at the molecular level is described, with a focus on the transforming growth factor-ß signaling molecule GDF11. Second, I explain how the cell population in the limb bud deforms with developmental progress, shaping the limb bud with elongation along the proximal-distal axis. Finally, I describe the developmental mechanisms that determine digit identity through the interdigits.