The linear dependence of the degree of polymerization on time confirms the conduct of radical polymerization in a controlled mode. The VT conversion was over 98% and the PVT number average molecular weight ranged from 11 to 61 kDa. The polydispersity of the synthesized polymers reached 1.16. The occurrence of the controlled radical polymerization was confirmed by monitoring the degree of polymerization over time.Narrow dispersed poly (PVT) was synthesized by reversible addition–fragmentation chain transfer (RAFT) polymerization of 1-vinyl-1,2,4-triazole (VT). AIBN as the initiator and dithiocarbamates, xanthates, and trithiocarbonates as the chain transfer agents (CTA) were used. Dithiocarbamates proved to be the most efficient in VT polymerization. Gel permeation chromatography was used to determine the molecular weight distribution and polydispersity of the synthesized polymers. The presence of the CTA stabilizing and leaving groups in the PVT was confirmed by  The datCarrying out the controlled radical polymerization of VT, in contrast to free radical polymerization, led to a decrease in the average molecular weights and the formation of narrowly dispersed polymers . The higThe effect of the natures of the RAFT agent, solvent, molar ratio of the initiator AIBN and CTA on the monomer conversion, molecular weight, and polydispersity of the resulting poly were studied .The use of methanol and DMF was due to the good solubility of the PVT and RAFT agents in these solvents. The molar ratio of [AIBN]:[CTA] ranged from 1:1 to 1:10. Regardless of the CTA used and the polymerization conditions, the polymer conversion was more than 98%, and the average molecular weight of the PVT varied in the range from 11 to 61 kDa . The useIn order to control the molecular weight of PVT, studies were carried out at different ratios of [M]:[CTA] in the range from 100 to 400 at a constant ratio of [AIBN]:[CTA] = 1:2.5 . Accordi2C≡N), so the efficiency of the reinitiation step for both chain transfer agents is close. The difference between CTA1 and CTA2 lies in the stabilizing group (Z), which ensures the stabilization of the macromolecular chain in the inactive (sleeping) state. Based on this, the difference in the molecular weight and dispersion of PVT is due to the influence of this fragment.The obtained results indicate that CTA2 is a more efficient RAFT agent in the controlled radical polymerization of VT compared to CTA1. CTA1 and CTA2 have the same leaving group  bond and easier attachment of the growing macroradical to polyCTA2. As a result, polyCTA2 had higher activity and a higher transfer coefficient compared to polyCTA1, which led to a decrease in the growth reaction rates and a decrease in M1H and 13C NMR spectroscopy. The 1H NMR spectrum of the poly obtained at [AIBN]:[CTA2] = 1:10 in DMF is shown in 1H NMR spectrum of the homopolymer. New broadened signals showed up in the region of 4.20–3.34 and 2.34–1.94 ppm, which were assigned to the proton resonance in the methyne and methylene groups in the polymer backbone, respectively. The signals at 7.87–7.29 ppm belonged to the protons of the triazole ring. In addition, the 1H NMR spectrum contained signals at 8.37–8.14 and 6.94 ppm, which were related to the protons of the pyridine ring of CTA2. The methyl proton signals of the dithiocarbamate group of CTA2 were visible at 3.80 ppm. The signal attributable to the methylene protons of the NC–CH2 R-group was detected at 2.43 ppm. Such signals were not observed in the 1H NMR spectrum of the PVT synthesized by free radical polymerization  and indicate that the polymer chain end is capped by CTA fragments as expected, according to the general mechanism of the RAFT process.Kinetic studies of VT radical polymerization in the presence of [AIBN]:[CTA2] = 1:2 in DMF at 60 °C were carried out in order to confirm the controlled nature of the polymerization. The conversion and molecular weight characteristics of PVT from the time of polymerization were controlled. The conversion, average molecular weight, and polydispersity of the homopolymer in the reaction mixture were determined by GPC.Monomer conversion and the ratio of the monomer concentration over time as a function of reaction time is shown in The dependence of the average molecular weight and polydispersity on the monomer conversion is shown in An important criterion for proving the controlled nature of polymerization is the ability to further increase the polymer chain, which is in an inactive state. For this purpose, the resulting and isolated polymer PVT , 16 was 1H and 13C NMR experiments revealed that the PVT chains contain CTA functional groups. The retention of activity of the PVT to chain transfer synthesized by the RAFT polymerization method was confirmed by successful chain extension.The controlled radical polymerization of 1-vinyl-1,2,4-triazole in the presence of AIBN as an initiator and dithiocarbamates, xanthates, and trithiocarbonates as the chain transfer agents was studied. Dithiocarbamates—cyanomethyl methyl(phenyl)dithiocarbamate and cyanomethyl methyl(pyridine-4-yl)dithiocarbamate—proved to be the most effective chain transfer agents. The use of methanol as a medium compared to DMF led to polymers with a narrower polydispersity. The study of RAFT polymerization at various [M]:[CTA] ratios indicates a linear dependence of the molecular weight, which confirms the controlled mode of VT polymerization. Regardless of the polymerization conditions, the monomer conversion was greater than 98%. The average molecular weight of the synthesized PVT was in the range from 61,000 to 11,000 Da while the polydispersity reached 1.16. The structural analysis performed via the


Caustic ingestion remains a complex public health issue worldwide, in adults and children. Caustic agent  is a product that causes tissue damage, and its ingestion will damage the mouth, pharynx, hypopharynx, esophagus, stomach, duodenum, and may reach the jejunum. Concomitant gastric and intestinal injuries can occur in 20 to 60% –3Acid products, most of the times, cause coagulative necrosis of the mucosa, while alkalis cause a liquefaction/saponification effect on the mucosa, reaching the entire wall of the organ. The most accessible acids are hydrochloric, sulfuric and oxalic acid, widely used in civil construction and factories. Products that contain alkali are those used in household cleaning, containing sodium hydroxide, also known as caustic soda. Therefore, caustic stenosis of the esophagus depends on the product ingested, the amount that was ingested and its concentration , 4–6.Ingestion of these products damages the esophagus, in different extensions, and the final consequence is progression to stenosis and fibrosis of the organ. The most common symptom is dysphagia, and the intensity of dysphagia depends on the degree of stenosis. Odynophagia is also present. The nutritional status injury is evident, due to significant weight loss , 2, 4, 7Developing countries have a higher incidence of this kind of accident. In children under 10 years of age, ingestion is most often accidental, however, in adolescents and adults, the etiology of ingestion is due to attempted suicide. In the United States, despite the decline in caustic intake in children , the incidence is 15.8 cases per 100,000 peoples .The treatment of esophageal strictures secondary to the ingestion of caustic products will depend on the extent of esophageal and gastrointestinal involvement, the degree of dysphagia caused to the patient and the clinical, systemic, nutritional repercussions and comorbidities. It is also important to assess gastric involvement, because gastric mucosal injury can lead to antropyloric stenosis. Zargar et al. observed that acute gastric injury was present in 85.4% of their patients who ingested acid, mainly involving the distal half of the stomach, with 44.4% presenting late complications, like pyloric or antral stenosis , 8.The first treatment attempt to relieve dysphagia is esophageal dilatation with the aid of digestive endoscopy, which can be repeatedly performed , 4, 9. IThe objective is to record the importance of continuous follow-up of these patients, the likely incidence of squamous cell carcinoma, the main known risk factors, treatment options and survival. In addition, the authors show the experience of the Service, in the follow-up of patients with previous ingestion of caustic products, in the last 40 years.Esophageal cancer is considered the seventh most common malignant tumor, with more than 570,000 new cases reported in 2018. Among the risk factors for esophageal squamous cell carcinoma are smoking, alcohol consumption, achalasia, drinking hot drinks, deficiencies in zinc, vitamins C, E and folates, consumption of red meat, socioeconomic status and genetic factors. Esophageal adenocarcinoma is associated with reflux esophagitis, Barrett’s esophagus, obesity and alcoholism .The ingestion of caustic products is a risk factor for the occurrence of esophageal cancer, and the literature reports the occurrence of the disease, approximately 30 to 40 years after the accident, mainly squamous cell carcinoma –18.The first description in the literature of the association of esophageal cancer in a patient with ingestion of caustic product was made in the literature by Teleky, in 1904 , 16, 17.The estimated risk in patients with a history of caustic ingestion is 1000-3000 times higher when compared to normal individuals of the same age group , 16–18. Usually, the risk of developing cancer is 2 to 30%, between 10 and 30 years after ingestion. Alcohol abuse and smoking should be risk factors to be considered in these patients , 16, 17.The occurrence of cancer is in the areas of narrowing and strictures in the esophagus, therefore, it is in these places that endoscopists, during the exams, need to focus their attention, in order to search for pre-neoplastic lesions , 9, 16The survival rate of patients with esophageal squamous cell carcinoma depends on the staging at the time of diagnosis and oncological and surgical treatment options , 19. HowRuol et al. analyzed 25 patients with esophageal scar cancer as late complication of caustic ingestion. The squamous cell carcinoma was diagnosed in 20 (80%) patients, adenocarcinoma in three (12%) and verrucous carcinoma in two patients. Esophagectomy was performed in 17 patients. The most frequent age of occurrence of carcinoma was between 40 and 70 years, with a median of 59 years .de Oliveira Junior et al. recorded the differentiated expression of miRNAs (miR-374 and miR-574) in esophageal mucosal biopsies of children with caustic strictures younger than 5 years, after the accident. The authors conclude that biomarker identification is a promising strategy to improve early diagnosis of esophageal cancer in caustic lesions that are at increased risk of progression .Tustumi et al. performed a systematic review analyzing the risk of malignant neoplasm of the esophagus and patients undergoing esophagectomy or esophagoplasty. The authors concluded that the latency period for cancer onset ranged from 22 to 58 years and the risk of cancer in patients with caustic strictures is 701.7 - 874.1 per 1,000,000 person-years .The digestive endoscopy must be performed, in the acute phase of ingestion of the caustic product, within the first 24 hours after the accident, in order to assess the lesion and depth extension. The Zargar classification is the most employed .The endoscopic follow-up is essential in the chronic phase of the disease. There is no consensus in the literature, on the minimum time interval between endoscopic exams, however, some authors recommend starting regular follow-up about 10-20 years after the caustic accident. They suggest that endoscopic surveillance be performed every 2–3 years, but the exact intervals are unknown , 25–27Pre-cancerous dysplastic lesions are detectable through digestive endoscopy and biopsies of suspicious areas. However, routine screening is currently not recommended outside high-risk regions or for low-risk individuals. Endoscopy remains the gold standard for diagnosing dysplasia and early squamous cell carcinoma, but it is an invasive and expensive method for the health system , 27, 28.Pennachi et al. employed Lugol’s iodine chromoendoscopy versus Narrow Band Image enhanced endoscopy to perform biopsies in suspected areas of 38 patients with caustic stenosis for early detection of esophageal carcinomas. There were 14 confirmed lesions detected with Lugol´s solution chromendoscopy and 9 with Narrow Band Imaging. All the suspected lesions were found adjacent to stenosis. The authors concluded that the general acuity of the exams was 73% .Eskander et al. analyzing the endoscopic biopsies of 100 children with caustic strictures undergoing endoscopic dilatations, of both sexes, with a mean age of 5.9 years, demonstrates evidence of chronic oesophagitis in 85%, 13% of reactive atypia in the form of severe neutrophilic inflammatory atypia and mild squamous dysplasia was diagnosed in two cases .When there is already associated carcinoma, staging is necessary and the exam indicated is a chest and abdominal computed tomography (CT), to evaluate the relationship between the esophagus and the airways, and the abdominal cavity. MRI can be used, but there are no advantages. Bronchoscopy is indicated to assess invasion of the trachea and bronchi , 28–30.Noh et al. analyzed chest CT scans of 14 patients with caustic strictures associated with malignant neoplasms that appeared on average 42 years after the caustic accident. The most common findings were eccentric wall thickening (71.4%), homogeneous esophageal wall enhancement (69.2%), periesophageal infiltration (78.5%) and enlarged mediastinal or hilar lymph nodes (14.3%) .Colonoscopy should be performed when the colon is considered as an option for by-pass such as esophagocoloplasty, especially in elderly patients, who may have intestinal polyps, tumors or diverticula , 8, 12The treatment options for patients with squamous cell carcinoma are chemotherapy (QTX), radiotherapy (RTX), chemoradiotherapy (CRT), immunotherapy, targeted therapies, endoscopic resection in early lesions and surgical treatment. Multidisciplinary or multimodal treatment is the most indicated, associating chemotherapy and radiotherapy alone, as neoadjuvants or adjuvants , 31. ImmTargeted therapy options are few, mainly employing targeting epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), or phosphoinositide 3-kinase/mammalian target of rapamycin (PI3K/mTOR) .The surgical treatment of the association of caustic stenosis and malignant neoplasm aims at resection of the lesion, performing esophagectomy. However, the literature records a reduced percentage of esophagectomies, due to advanced disease at the time of diagnosis Table 1.The number of esophagectomies performed is small, it was not possible to assess either the authors’ preference for the access route for esophagectomy , nor the most used reconstruction , 32, 33.The need or not of resection of the ill esophagus, during the reconstruction of the digestive tract, is discussed in the literature, due to the occurrence of malignant neoplasm in the dysfunctionalized organ in the mestiastinum and difficult access and subsequent diagnosis. These same authors report postoperative complications at significative rates, resulting from esophagectomy associated with bypass , 32.The malignant neoplasms in patients with a history of ingestion of caustic products is a real fact and has been recorded 30 to 40 years after the accident.There is no gender preference and no more prevalent age. Tabagism and alcoholism increase the risk of cancer.Squamous cell carcinoma is the most prevalent and occurs in areas of strictures.The diagnosis of neoplasia is difficult and periodic endoscopic follow-up, with biopsies, is necessary for all patients with a history of caustic ingestion.Biopsies should be obtained close to areas of strictures, where neoplasms arise.The unacceptably high incidence of caustic ingestion in some countries and regions highlights the need to implement prevention programs and continuing adult education.All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Kallmann syndrome (KS) is a rare hereditary disease with high phenotypic and genetic heterogeneity. Congenital hypogonadotropic hypogonadism and hyposmia/anosmia are the two major characterized phenotypes of KS. Besides, mirror movements, dental agenesis, digital bone abnormalities, unilateral renal agenesis, midline facial defects, hearing loss, and eye movement abnormalities can also be observed in KS patients. Because of the phenotypic heterogeneity, genetic diagnosis become increasingly valuable to distinguish KS from other disorders including normosmic congenital hypogonadotropic hypogonadism, constitutional delay of growth and puberty, CHARGE syndrome, and functional hypogonadotropic hypogonadism. Application of next-generation sequencing has promoted the discovery of novel pathogenic genes in KS pedigrees. Prenatal diagnosis is an effective method in clinical settings to decrease birth defects and block transmission of genetic disorders. However, pregnant women may suffer from physical and psychological distress when fetuses are diagnosed with congenital defects. Preimplantation genetic testing (PGT) is a prospective approach during the in vitro fertilization process that helps to interrupt transmission of hereditary diseases to offspring at an early stage. Thus, genetic testing and counseling are recommended to KS patients with family histories, prenatal diagnosis and PGT are considered to be useful options. Kallmann syndrome (KS) is a hereditary disease characterized by congenital hypogonadotropic hypogonadism (CHH) and hyposmia or anosmia. Isolated gonadotropin-releasing hormone (GnRH) deficiency (IGD) is a common disease with genetic and clinical heterogeneity, and KS is the main component of IGD. The estimated incidence of KS in males 1:30,000) is 4–5 times higher than in females 1:125,000), but the latter is probably underestimated because affected women tend to have milder symptoms than affected men [:30,000 i5,000, buCHH and hyposmia or anosmia are the two typical symptoms of KS. The correlation between anosmia and small testes in individuals was first noted by a Spanish pathologist, Aureliano Maestre de San Juan, in 1856 . In 1944GnRH or luteinizing hormone-releasing hormone (LHRH), a decapeptide, is essential for mammalian puberty onset and reproduction. The reproductive function is mainly managed by 1200–1500 GnRH neurons, and these unique neurons have either an olfactory placode/ectodermal or a neural crest cell origin –8. GnRH In 1989, Schwanzel-Fukuda et al.  and WrayThe hypothalamic-pituitary-gonadal (HPG) axis is activated from the mid-trimester, restrained at the end of gestation, and reactivated after birth. In early childhood , GnRH pulsatile secretion is actively inhibited and persists until adolescence . ReactivNeonatal period and childhood In male infants, cryptorchidism and micropenis (stretched penile length <2.5 cm) [Adolescence Because of their low GnRH levels, KS patients show absent or partial pubertal development in adolescence. In male adolescents, absent and/or minimal virilization, low libido, and lack of sexual function are the common symptoms [Adulthood KS diagnosis in adulthood is mainly associated with infertility and sometimes with early appearance of osteoporotic fractures [CHH and hyposmia or anosmia are the most typical characteristics of KS. Because HPG axis activity and function fluctuate during the lifespan, the axis provides a good time-window for diagnosis of KS. Based on the influence of GnRH participation in different periods of growth and development in humans, the symptoms of KS can be summarized into three stages: (1) neonatal period and childhood, (2) adolescence, and (3) adulthood. <2.5 cm) , 19 are <2.5 cm) . In GnRHsymptoms . Absencesymptoms . Adulthoractures , 21. SomBesides the symptoms mentioned above, small numbers of KS patients show synkinesia (mirror movements), dental agenesis, digital bone abnormalities , unilateral renal agenesis or urinary tract malformation, midline facial defects , hearing loss, eye movement abnormalities, poor balance through cerebellar ataxia, skeletal anomalies , and pigmentation defects –25.The psychological problems of KS patients are non-ignorable, especially among male adolescents, and include depression, anxiety, and decreased quality of life that leads to low self-esteem and other effects . CompareIn the first several months after birth, transient activation of the HPG axis is defined as “mini-puberty” . During The clinical diagnosis of KS includes evaluation of CHH and hyposmia or anosmia. The diagnosis of KS can rely on reproductive symptoms, such as history of cryptorchidism and/or micropenis , presencHormonal analysis provides useful indicators for KS diagnosis. Male and female KS patients both have very low plasma gonadotrophin, FSH, LH, and inhibin B levels. During mini-puberty in particular, serum levels of testosterone  and FSH and LH  show transient increases that are extremely low in infants with CHH .Meanwhile, some brain changes have been noted in KS patients. Magnetic resonance imaging examination can show abnormalities in the olfactory fossae, such as unilateral or bilateral olfactory bulb or tract agenesis, reduced depth, reduced curvature, or increased cortical thickness within the olfactory sulcus in KS patients , 32, 33.Although the majority of KS cases are sporadic, many cases are clearly familial, and more than 30 mutated genes in KS patients have been identified to date . These gANOS1 , the first and most common pathogenic gene for KS identified in 1991, is inherited through the X-linked recessive mode [ANOS1, located on Xp22.3 and consisting of 14 exons, encodes the protein anosmin-1, a transient and regionally restricted extracellular adhesion protein that participates in GnRH neuron adhesion and axonal migration during organogenesis [ANOS1 and cause frameshifts or premature termination codons during protein translation process [ANOS1 showed disconnection of olfactory, vomeronasal, and terminalis nerve fibers with the brain, and GnRH neurons were unable to migrate to their normal position in the brain [ANOS1 mutations are found in 10%–20% of all KS patients, including 30%–60% of familial cases and 10%–15% of sporadic cases [ive mode . ANOS1, ogenesis , 35. Anoogenesis . Nucleot process . A 19-wehe brain , 36. ANOic cases –40.FGFR1 was reported to be mutated in KS families by Dodé et al. in 2003, and FGFR1 mutations are the second most common gene mutations found in KS patients. FGFR1 encodes fibroblast growth factor receptor 1 (FGFR1), which belongs to the receptor tyrosine kinase superfamily [FGFR1 mutations in KS patients is ~10% [FGFR1 is located on chromosome 8p11.23 and follows the AD/AR/oligogenic inheritance modes. FGFR1 consists of an extracellular domain (three immunoglobulin-like domains that determine the affinity and specificity for its ligands), a transmembrane helix domain, and an intracellular tyrosine kinase domain [FGFR1 mutations include missense, nonsense, splice site, frameshift mutations and intragenic insertions or deletions, and the phenotypes arising from these different mutations are heterogeneous [FGFR1 mutations are associated with delayed puberty, anosmia, cleft palate, mirror movements, dental agenesis, and bimanual synkinesia, although some familial cases have males and females with FGFR1 mutations who are asymptomatic carriers [FGFR1 mutation was reported to show a reversible condition after several years of testosterone treatment, and the same mutation was found in his mother and maternal grandfather [FGFR1 mutation dominant and recessive forms in KS remain unclear.erfamily , 42. The is ~10% . FGFR1 ie domain . FGFR1 mogeneous –45. FGFRcarriers –48. Thercarriers . SurprisPROKR2 and PROK2 encode the G-protein-coupled receptor prokineticin receptor 2 (PROKR2) and its ligand prokineticin-2 (PROK2), respectively, and the PROKR2/PROK2 pathway plays essential roles during olfactory bulb development and GnRH neuronal progenitor migration and differentiation [PROKR2 and PROK2 mutations can be heterozygous, homozygous, or compound heterozygous, and the prevalence of these mutations in KS patients is about 5%–10% [Prokr2 or Prok2 knockout mice were found to show hypoplasia of the olfactory bulb and reproductive system, without the normal architecture [ntiation . PROKR2 t 5%–10% , 50. Proitecture , 52. KS itecture .CHD7 located on chromosome 8q12 consists of 38 exons, and encodes chromodomain helicase DNA-binding protein 7 (CHD7). Mutations of CHD7 were identified in patients with CHARGE syndrome, KS, and CHH [CHD7 was originally considered to be the causative gene for CHARGE syndrome, subsequent research showed that CHD7 mutations can also be detected in KS patients lacking mutations in KAL1, FGFR1, PROK2, and PROKR2 [CHD7 mutations in all KS/nIHH patients is predicted to be >5% [ and CHH . Althougd PROKR2 . The preo be >5% , 57.FGF8/FGFR1, SOX2, CHD7, FGF17, and IL17RD are involved in the GnRH neuron fate specification process [ANOS1, PROK2/PROKR2, SEMA3A/PLXNA1, SEMA3E, NSMF (NELF), HS6ST1, WDR11, SOX10, FEZF1, IGSF10, DCC/NTN1, TUBB3, and SMCHD1 are associated with olfactory axon guidance, GnRH neuron migration, or axon projection [As well as the major genes described above, the development of NGS has led to the identification of many other pathogenic genes in KS patients during the last several decades. The majority of the mutated genes identified to date participate in GnRH neuron development and migration or secretion processes.  process , while Aojection  (Table 1PROKR2 (p.L173R) and ANOS1 (p.S396L) [In addition to the classical Mendelian inheritance pattern, digenic and oligogenic modes have been described in KS patients . First mp.S396L) . ApplicaDespite the diagnostic criteria mentioned above, the key point for KS diagnosis is to clarify the differential diagnoses. The main differential diagnoses of KS include tumors caused by acquired hypogonadotropic hypogonadism, constitutional delay of growth and puberty (CDGP), CHARGE syndrome, and functional hypogonadotropic hypogonadism , 60, 61.FGFR1, FGF8, FGF17, NSMF, SPRY4, IL17RD, DUSP6, GLCE, FLRT3, PROK2/PROKR2, HS6ST1, HESX1, CHD7, KLB, PLXNA1, WDR11, DCC, NTN1, SEMA7A, and SEMA3E [OTUD4, RNF216, POLR3A, POLR3B, PNPLA6, PITX2, STUB1, SOX2, SOX3, OTX2, PROP1, PCSK1, LHB, FSHB, LHX4, LHX3, DMXL2, GNRH1, GNRHR, GLI2, GATA2, IGSF10, KISS1/KISSR1, TAC3/TACR3, LEP/LEPR, TBX3, NR0B1, and CCDC141 gene mutations were only reported in nCHH [Normosmic congenital hypogonadotropic hypogonadism (nCHH) shares similar phenotypes with KS, including bimanual synkinesia, renal anomalies, facial cleft, dental agenesis, eye movement disorder, and hearing loss, although the prevalences of these abnormalities are more common in KS than in nCHH , 64. Thud SEMA3E . Besides in nCHH . The comGNRHR, FGF8, and PROKR2 [Adult-onset hypogonadotropic hypogonadism (AHH) occurs in healthy adult males who have completed normal puberty and have proven fertility . AHH cand PROKR2 , which aColoboma, Heart defect, Atresia of choanae, Retardation of growth and/or development, Genital hypoplasia, and Ear anomalies [CHD7 are identified in >75% of patients with CHARGE syndrome. KS and nIHH were hypothesized to be milder allelic variants of CHARGE syndrome [CHARGE syndrome is a multisystem disorder that includes symptoms of nomalies . CHARGE nomalies . Mutatiosyndrome . KS and syndrome , 70. TheEarly diagnosis and treatment of KS are crucial, because they can lead to improved quality of life and prevention of disease-related complications, especially during adolescence as the vital period for secondary sexual maturation and functioning and the critical point for adolescent psychological development. As mentioned before, CHH and hyposmia/anosmia are the two main symptoms of KS, especially CHH causes the most disturbed symptom in KS patients’ life. Surgical operative is the first choice for newborns with cryptorchidism at 6–12 months. HRT promotes virilization, development of muscle mass and strength, pubertal growth spurt, deepening of voice, growth of penis, libido, and sexual function in males  and furtAlthough current treatments markedly improve quality of life in KS patients, the genetic characteristics of KS mean that there is some risk of transmitting the mutated genes to offspring and presenting severe symptoms.Prenatal screening/diagnosis is an effective method in clinical settings to decrease birth defects and block transmission of genetic disorders. Different methods can be used, including maternal serological screening markers, fetal chromosome karyotype analysis, fetal ultrasound, and non-invasive prenatal testing . KS fetuAlthough the majority of infertile male and female patients with KS have a good reproductive prognosis after appropriate HRT or through assisted reproductive technologies (ART), all KS patients who seek fertility should receive counseling and discuss with their doctors or genetic specialists about the risk of transmitting the disease to their offspring , 58, 77.In recent years, besides Sanger sequencing and other traditional methods used for PGT, the booming development of high-throughput NGS technology had led to its common use in PGT, providing improved resolution and increased genetic information and reducing the cost and time required for sequencing . MultiplIn 2019, Chan et al. reported the first live birth of a term baby girl delivered into a family wherein the husband had KS with anosmia and absence of secondary sexual characteristics . An FGFRFGFR1 mutation was found in both the patient and his father. However, the father did not show any symptoms of KS, which may be related to incomplete penetrance and variable expressivity among different individuals or oligogenicity. For this family, whole-exon sequencing would be required to identify other potential pathogenic genes. Thus far, genetic causes can be determined in 50–60% of all KS cases, which is vital for diagnosis and fertility prognosis of KS patients, and PGT has become an optional approach to block transmission among generations.In this reported case, only 30 KS- or CHH-related genes were tested, and the With the current improvements in its efficiency and security, PGT has become a useful approach for familial inherited diseases to prevent transmission of pathogenic genes to the following generation. However, thus far, KS patients have very rarely chosen to acquire their babies through PGT procedures in clinical settings. This may arise because the majority of male KS patients can produce sperm and acquire offspring after a regular period of HRT, the symptoms are mild in some KS patients, or there is economic pressure. The majority of KS patients choose to acquire their offspring by natural fertilization or IVF after HRT. For patients with family histories, genetic testing and counseling are recommended to establish the mutated genes, associations between pathogenic genes and phenotypes, and inheritance patterns. In families with severe inherited symptoms, such as congenital heart defect, hearing loss, and cleft lip/palate, PGT can be an optional approach to avoid offspring suffering from KS. However, in some cases, there are some limitations and challenges for genetic counseling caused by incomplete penetrance, and oligogenicity for known and unknown genes. Therefore, PGT is a perspective genetic interruption method for KS. Both methods have their advantages and disadvantages, so that it is difficult to evaluate which one is obviously better than the other. For PND, fetal samples are obtained by invasive methods  for genetic prenatal diagnosis to find whether the fetus carries genetic abnormalities. But the pregnant woman may suffer from psychological stress if the fetus has been diagnosed with KS. While, PGT can avoid such a situation by selecting and transfer an embryo without genetic abnormalities, but OHSS may occur during PGT procedures, and the biopsy procedure is invasive and may be harmful to the embryos.The current clinical management and genetic interruption perspective of KS are summarized in Fig. In conclusion, KS is a rare hereditary disease with high phenotypic and genetic heterogeneity. In the last decade, pathogenic genes have gradually been identified in KS patients. Although appropriate HRT or ART can help infertile male and female patients to achieve a good reproductive prognosis, their offspring still have risks of suffering from KS. Although a prenatal diagnosis can detect congenital defects during pregnancy, pregnant women may suffer from physical and psychological distress, especially when the fetus is found to be carrying genetic defects. Meanwhile, with the advances and application of NGS and single-cell amplification technologies, as well as improvements in safety and accuracy, PGT has helped thousands of families with hereditary diseases to obtain a healthy next generation. Although no successful case with genetic interruption of PGT to KS patients was reported until now, genetic counseling is an essential component for KS patients who seek fertility, and it can be an imaginable outlook that PGT becomes an optional approach in the near future to decrease the risk of transmitting the disease of KS to their offspring.


The development of a dismantlable adhesion technologythat allowsswitching between bonding and debonding states using external stimuliis important for realizing renewable and sustainable material cycles.Controlling the adhesion interface is an effective approach to manipulatethe adhesion strength; however, research on dismantlable systems focusingon the interface has not been proceeded. Recently, we demonstrateda novel dismantlable system based on a stimuli-responsive molecularlayer comprising cleavable anthracene dimers, which strengthen theinitial adhesive force by forming chemical bonds between the substrateand adhesive and can be dismantled when required via stimulation-inducedbond breaking. Here, we evaluate the use of the anthracene-based molecularlayer with different components for verifying its versatility in theadhesive/dismantling system. The formation of the cleavable molecularlayer by the stacking of relevant molecules enabled its usage withtwo types of adhesives, an epoxy adhesive and a silane-modified polymeradhesive. The initial adhesive strengths were improved in both typesof molecular layers by creating chemical bonds at the adhesion interfaces.Light irradiation or heating stimuli for 1 min reduced the peel strengthby up to 65%, and dismantling occurred in the cleavable photodimerlayer. This study expands the versatile applicability of the molecularlayer-based dismantling system. The debonding mechanisms of dismantlable adhesives are mainlybased on changes in the bulk properties of the adhesive, making itdifficult to achieve both strong bonding and easy debonding by controllingthe cohesiveness of the adhesive.Adhesion is an important process in theproduction of a range ofproducts using bonding materials.21 One typical process involvessurface treatment with a primer, which has been conventionally usedto efficiently improve adhesion strength between materials.24 The related studies have indicated that thin layers formed at amolecular level by coating processes can influence adhesion strength.At the adhesion interface, physical or chemical bonds  have been suspected to play an importantrole in high-strength adhesion because of the strong bonding providedby such linkages.1 Hence, it is possibleto realize both strong adhesion and easy separation by switching thechemical bonding at the adhesion interface.26 However, because of the difficulty in controlling the formationand breakage of chemical bonds, the progress in studies aimed at changingthe chemical bonding state at the interface to reduce adhesion strengthhas been slow.Another approach to controlthe adhesion strength is to treat thesurface of the adherend with a focus on the state of the adhesioninterface.28 In the molecular layer, the chemical bonding was controlled usinganthracene molecules, which are known to undergo photodimerizationand thermal/photocleavage reactions under external stimuli.30 When heat or light is applied to the cleavable molecular layer formedat the interface between the substrate and adhesive, the adhesionstrength decreases, and separation occurs at the molecular layer.28 In addition, this molecular layer not only functionsas a dismantlable system but also acts as a primer that increasesthe adhesion strength in the initial state. This function is achievedby the chemical bond formation between the molecular layer and thereactive functional groups of both the substrate and the adhesive.Thus, designing reactive functional groups in the molecular layeris important to realize the concept of this system.We previously developed a system that could bedismantled fromthe adhesion interface using a stimuli-responsive molecular layerconsisting of chemical bonds that could be cleaved by applying a stimulus.31 This processenables changes in the molecular layer composition according to thedesired purpose, such as the presence of cleavable units and the selectivityof the functional group at the outermost surface. In this study, weinvestigated the effect of the composition of the molecular layeron a molecular layer-based dismantling system to explore the versatilityof this system. Using molecular layers with or without the cleavableunit of the anthracene photodimer, we verified the concept of thedismantling system based on the chemical bond change. In addition,the functional group at the outermost surface of the molecular layerwas changed by controlling the stacking procedure of 9-anthracenecarboxylicacid dimer (Di9AC) and 3-aminopropyltriethoxysilane (APS). We preparedtwo types of molecular layers containing carboxyl groups or triethoxysilylgroups at the outermost surface, which enables its reaction with differenttypes of adhesive components  and APSwere procured from Tokyo Chemical Industry Co., Ltd. Other reagentsand solvents were purchased from Tokyo Chemical Industry Co., Ltd.,Wako Pure Chemical Industry Ltd., or Kanto Chemical Co., Inc. andused without further purification unless otherwise stated. Di9AC wassynthesized according to a published procedure.2 pieces.27 The cut quartz substrates were cleaned ultrasonically intwo consecutive steps using a 2.0% neutral detergent (Extran MA-02)solution in water and pure deionized water .Subsequently, they were immersed in an alkali detergent consistingof a 10 wt % solution of KOH  in ethanol for 1 d. Thesubstrates were then rinsed four times with deionized water throughsonication for 20 min. Before use, the cleaned substrates were driedunder vacuum for 1 h to ensure the removal of the residual adsorbedwater.Quartz substrates purchasedfrom iTEC Science Co. Ltd. were cleaned as reported previously aftercutting them into 30 × 10 mm31 First, the cleaned andcompletely dried substrates were immersed in an APS solution  for 1 h at 25 °C for aminosilylation. They weresubsequently washed with dry toluene, heated at 100 °C for 30min, sonicated in toluene for 1 min, and then dried under ambientconditions to obtain the substrate covered with the molecular layerconsisting of APS molecules (APS layer). Next, the aminosilylatedsubstrates were immersed in a Di9AC solution for 30 min at 40 °C in a shaking bath. Thereafter, the substrateswere removed gently and dried for 30 min at 80 °C. The substratescovered with the molecular layer consisting of APS and Di9AC (APS/Di9AClayer) were finally rinsed with acetone and dried under ambient conditions.Substrates covered with a molecular layer of 9AC (APS/9AC) were preparedusing a similar process. For stacking APS molecules onto the Di9ACmolecules, the substrates covered with the APS/Di9AC layer were immersedin an APS solution  for 30 min at 40 °Cin a shaking bath, removed gently, and dried for 30 min at 80 °C.Finally, the substrates with the stacked molecular layer (APS/Di9AC/APS)were rinsed with acetone and dried under ambient conditions.Dismantlable molecular layerswere formed on the cleaned quartz substrates according to a previouslyreported method.2.For thermal cleavageof the molecular layer, the substrate was heated using a Mettler FP82HT. Light irradiation for photocleavage was performedusing a light-emitting diode .The light intensity was controlled to be 10 mW/cmUltraviolet–visible(UV–vis) absorption spectra were measured to investigate theprogress of the cleavage reaction using a JASCO V-670 spectrometer.Fluorescence spectra were recorded in the solid-state configurationusing a JASCO FP-8550 spectrometer. Differential scanning calorimetry(DSC) was conducted at a heating and cooling rate of 10 °C/minusing a HITACHI DSC 7000X.2) with the deposited molecular layer and cleanedbare quartz substrates (without a molecular layer) were used in thisevaluation. The bare substrate was cleaned as described above. Thespecimens for peel strength measurements were prepared according toa previous report.28 Typically, the substratewas adhered to a hydrophilic polyester film  using an adhesive. Two types of adhesives were selecteddepending on the composition of the molecular layer. The thicknessof the adhesive layer was adjusted to be 100 μm. To completethe curing of the adhesive, the adhered specimens were maintainedat 25 °C in an environment with a humidity of >50% for 24h.After this process, a stimulus such as light irradiation or heatingwas applied as required from the quartz substrate side. The quartzsubstrates bonded with polyester films were adhered to a glass slideto fix them to the tensile test machine. The adhesion tests were performedat 25 °C using a tensile test machine  equipped with an accessory for 90° peel tests . The peeling rate in the 90° peel testwas 50 mm/min. The unit of peel strength was N/10 mm, based on thewidth of the specimens. The mean value of five replicate samples (n = 5) is reported as the peel strength.Quartz substrates (30× 10 mm34 We prepared two types of cleavable molecular layer(APS/Di9AC layer and APS/Di9AC/APS layer) on a quartz substrate bystacking molecules based on the layer-by-layer process. The aminogroups in APS units interacted with the carboxyl groups of the Di9AClayer via acid–base–type reaction . We determined that the anthracene dimer could be cleavedin a short duration by heating it at higher temperature. However,a decreasing trend in the absorbance due to the sublimation of theanthracene molecules is detected with continued heating, as the sametendency as in the previous study.31A cleavable molecular layer was formed on a quartz substrate by stackingfunctional molecules see 1, and thtion see 1b, leadi minutes 2a. Howev minutes 2b. This Figure S2). This implies that photoirradiation and heating of the APS/Di9AClayers helped the cleavage reaction to proceed, and the produced anthracenemonomers were stacked monomerically on the substrate.In addition, the photocleavage reaction was monitoredusing fluorescencespectroscopy. 24 The test specimen is prepared by adheringa quartz substrate and a flexible polyester film, as illustrated in To investigate the effect of the stimulation for causing the dismantlingsystem on the adhesion test, we conducted 90° peel tests usingbare substrates, which have no molecular layer. We conducted peeltests on the specimens composed of the substrate without molecularlayer to evaluate the effect of the stimuli on the bulk states ofthe adhesives. The peel strength is known to reflect the state ofthe adhesive/adherend interface, and a rigid specimen bonded witha flexible substrate on at least one side is used to measure the peelstrength.Figure S3, no significant difference in the peel strength was observed witha change in the stimulus. However, upon comparing the physical statesof the peeled specimens, the adhesive remained on the entire surfaceof the substrate only in the case of the heated sample. As the adhesivewas discolored, we inferred that heating at 180 °C degraded theadhesive component, even if the heating time was only 1 min. Therefore,we investigated the effect of heating at a lower temperature. Thethermal cleavage reaction in the molecular layer proceeds upon heatingfor a long time at a lower temperature of 140 °C, as shown in Figure S1; then, the specimen was heated at 140°C for 90 min. However, even in this case, the adhesive was discoloredand remained on the detached substrate . The DSC measurement of the epoxy adhesive implied thatthe heating stimuli induced the decomposition of the adhesive componentsbecause a significant peak was observed above 100 °C in the firstheating process . And also,the baseline shift was observed for both the heating and the coolingprocesses at 77.0 and 77.4 °C, respectively. These results indicatethe heating stimulus is not suitable for inducing dismantlable adhesionwith this epoxy adhesive.Based on the resultsof the cleavage reaction of the APS/Di9AC molecular layer 2, the ap40 Thus, it is reasonable that covalent bonds are createdat the adhesion interface by these molecular layers. Note that themolecular layer prepared by layer-by-layer deposition includes acid–baseinteraction between the stacked molecules. The bond energy of hydrogenbonds and covalent bonds are around 2.5–125 and 200–800kJ/mol, respectively.41 This implies thatthe acid–base interaction is weaker than the covalent bonds.In spite of this, we confirmed an improvement in the initial adhesivestrength in the case of using APS/Di9AC layer. Therefore, it is reasonablethat the bonding strength of the acid–base interaction is strongenough for this specimen.Figure S5). In contrast, in the case ofthe test specimen prepared with the APS/Di9AC layer, the peel strengthdecreased significantly after the light irradiation , and the failure modes of the stimulated specimensalso reflected the influence of the stimulation , the lightirradiation had no effect on the peel strength and the failure modesS5. In coConsidering the principle of the dismantlable system consistingof a cleavable molecular layer, anthracene monomers generated underthe applied stimuli are expected to remain on the quartz surface.Characteristic fluorescence bands of anthracene were detected by thespectroscopic measurement of the detached specimens after dismantlingby light irradiation 4c. This Figures S7 and S8), and the calculated resultsare summarized in S7), the failure modes were different for theexistence of the molecular layer: cohesive or interfacial failureoccurring for the specimens with or without molecular layer, respectively.These results are strong evidence that the molecular layer at theinterface affects the adhesion strength. Note that the compositionof the APS/Di9AC/APS layer is almost the same and uses the same polymeradhesive as that reported previously.28 The average peel strength of the test specimen of the APS/Di9AC/APSlayer was determined to be 10 N/10 mm, which is almost the same asthat observed in our previous report.28The cleavable molecular layer was implemented withdifferent functional groups, making it possible to form chemical bondswith different types of adhesives. Therefore, we designed the molecularlayer as an APS/Di9AC/APS layer, whose surface was covered by triethoxysilylgroups to react with the silyl groups of a silane-modified polymeradhesive. The peel tests of the specimens with and without the molecularlayer were conducted and tensile force plots were obtained S7 and S828 In this study, the coverage rate of the anthracene dimer,which is the key material responsible for dismantling, might be lowbecause of the stacking procedure. Particularly, in the case of applyingthermal stimuli, the specimen bonded tightly owing to the softeningof the adhesive under heating. Therefore, the difference in the coveragerate of molecules due to changes in the molecular layer formationprocess possibly causes a decrease in the dismantling ability as comparedwith the previous study.28The dismantling properties of the APS/Di9AC/APSlayer were evaluatedby applying a stimulus to the specimen. The conditions of the appliedstimuli were determined based on the cleavage characteristics of theanthracene dimer: irradiation with 280 nm light for 1 min or heatingat 180 °C for 1 min. While the test specimens consisting of asubstrate without a molecular layer showed the same peel strengthbefore and after the stimulation, the peel strength of the test specimencovered with the multimolecular layer was significantly decreasedin both cases of light irradiation and heating 5a. As coUpon comparingthe peel strength obtained using the epoxy adhesivesand the silane-modified polymer adhesives 4a and 5a45 and we successfully constructed the molecular layer by stackingappropriate functional molecules. We measured the peel strength ofthe substrate coated with the molecular layer and bonded to a flexibleadherend using two types of adhesives: an epoxy-based adhesive anda silane-modified polymer adhesive. The molecular layers were designedindividually for each adhesive, and in both cases, the strengtheningand weakening of the adhesive force at the interface could be controlledusing an external stimulus. One of the greatest strengths of thissystem is its ability to take advantage of the adhesive propertiesof functional adhesives such as the adhesion strength because thissystem only affects the adhesion interface. In fact, dismantling fromthe adhesion interface was observed regardless of the type of adhesiveused. Moreover, the peel strength of the test specimen in the initialstate reflected the cohesiveness of the adhesive. The results indicatethat the molecular layer-based dismantling process is highly attractivefor realizing renewable and sustainable material cycles.In this study, we investigated the performanceof an anthracene-basedcleavable molecular layer that facilitates strong adhesion and subsequenteasy separation at the adhesion interface when desired. The bondingand debonding could be achieved based on the state of the chemicalbonds at the adhesion interface. The concept of this multimolecularlayer system was inspired by the basic theory of the layer-by-layerassembly developed for functionalizing various surfaces,


It is vital that there are coordinated, collaborative efforts to address the threat of antimicrobial resistance (AMR) and to prevent and control the spread of hospital-onset infections, particularly those due to multidrug-resistant (MDR) pathogens. The butterfly effect is a concept in which metaphorically speaking, small, seemingly trivial events ultimately cascade into something of far greater consequence, more specifically by having a non-linear impact on very complex systems. In this regard, antimicrobial stewardship programs (ASP), when implemented alongside infection prevention control (IPC) interventions in hospitals, particularly hand hygiene (HH), are significantly more effective in reducing the development and spread of AMR bacteria than implementation of ASP alone. In this perspective, we briefly review the evidence for the combined effect, and call for closer collaboration between institutional IPC and ASP leadership, and for well-functioning IPC programs to ensure the effectiveness of ASP.  Escherichia coli, followed by Staphylococcus aureus, Klebsiella pneumoniae, Streptococcus pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa. The prevalence of infections caused by antimicrobial-resistant (AMR) bacteria is rapidly increasing and this, along with the constantly evolving global epidemiology, represents a major challenge . RecentlP. aeruginosa (CRPA), and carbapenem-resistant A. baumannii (CRAB), which are resistant to all routinely available antibiotics, are being reported [In this regard, increasing numbers of Gram-negative bacteria (GNB), such as the carbapenemase-producing Enterobacterales (CPE), carbapenem-resistant reported ,3. As sureported . Notablyreported .In response, there have been numerous calls for awareness and for strategies designed to attenuate these poor outcomes and to prevent the further development and spread of AMR. Major strides have been made in recent years, including the World Health Organization (WHO) approval of the Global Action Plan on Antimicrobial Resistance in 2015, and in 2019 AMR was declared one of the top 10 public health threats facing humanity [Compounding this challenge, is that whatever positive traction had been gained in combatting AMR during recent years, the SARS-CoV-2 pandemic has resulted in significant setbacks that have adversely affected progress ,6,7. ThiWhereas SARS-CoV-2 infection prevention and control (IPC) initiatives may have contributed to a reduction in AMR rates in some institutions, paradoxically, despite the adoption of strict COVID-19 IPC protocols, this did not occur in most hospitals and networks globally . MultiplS. aureus and methicillin-resistant S. aureus (MRSA) in this population and confirmed that MRSA was a common causative agent of pneumonia in patients with COVID-19 [K. pneumoniae (bloodstream infections), CRPA (ventilator-associated pneumonia), and CRAB . Significant heterogeneity was however apparent, mostly in the large, system-wide, regional, or national comparative assessments relative to single-centre studies [Concurrent bacterial and fungal infections have played an important role in hospital outcomes during the COVID-19 pandemic [COVID-19 . In termCOVID-19 . This wa studies . The imp studies . In this regard, Timsit et al. recently summarized the available evidence, emerging options, and unsolved controversies for the sequential optimization of antibiotic therapy in the intensive care unit (ICU), the site where collateral damage is most apparent . This naIn chaos theory, the butterfly effect describes the often-unintended consequence of small changes in the initial conditions of the state of a deterministic nonlinear system, which subsequently manifests as large differences. Conceptually, this small, seemingly trivial event may ultimately result in a non-linear impact on very complex systems . MathemaIn this regard, a number of reviews, which included publications from both high and low-income settings, have clearly demonstrated that ASPs, when implemented alongside IPC measures, and in particular measures to improve hand hygiene (HH) compliance, are significantly more effective in hospitals than the implementation of an ASP alone ,15,16,17As an example, Baur et al. reported that studies that co-implemented a HH intervention along with an ASP reduced AMR bacteria by 66% versus 17% in those that did not . The divHowever, data describing the impact of bespoke IPC and ASP interventions on colonization pressure and resistance rates of specific highly resistant bacteria such as extended-spectrum β-lactamase (ESBL)-producing Enterobacterales, CPE, CRPA, or CRAB, are scarce, and if performed may be more useful to direct and inform specific collaborative ASP and IPC interventions. In this regard, a systematic review and network meta-analysis of the relative efficacy of strategies for the prevention of AMR-GNB in the ICU provided some insights . In thisCompared with the STD arm, a multifaceted strategy comprising STD, ENV, SCT, and ASP was the most effective intervention to prevent MDR-GNB acquisition in the overall analysis, and for each specific type of bacteria . NotablyA. baumannii resistance and a sustained reduction in CRAB rates in a tertiary ICU in Beirut, Lebanon [In contrast, Rizk et al., utilizing existing resources in a low-income setting, recently demonstrated how a multi-disciplinary approach and combined interventions by both ASP and IPC teams led to an overall decrease in  Lebanon . This inClostridioides difficile and carbapenem-resistant Klebsiella pneumoniae (CRKP) infections [C. difficile appeared to focus more on stewardship, while those targeting CRKP focused more on screening, isolation precautions, or environmental disinfection as core strategies. Whilst some studies that incorporated multifaceted interventions, including HH or other IPC programs and ASP interventions concurrently, were shown to reduce HOIs, improve the rational use of antibiotics, and reduce mortality, there was limited evidence as to how the interventions influenced compliance with any of the interventions. In this regard, a commitment, and a call to strengthen and expand qualitative research efforts to improve the impact on compliance by ASP and IPC programmes, and the impact on AMR, is vital [A scoping review recently assessed ASP and IPC interventions targeting healthcare-associated fections . Notablyis vital . In addiThe data presented provide overall support for the integration of IPC practitioners into the ASP teams and the necessity of promoting adherence to IPC measures, including the level of HH compliance, prior to the development of an ASP that mostly relies solely on antibiotic processes and outcome measures. ,14. The Firstly, to ensure a hospital-wide culture that promotes behavioral change through recognition that by reducing AMR bacteria together, patient safety is improved.Secondly, to neutralize and amalgamate the clinical, microbiology, and IPC operational ‘silos’ within institutions.Thirdly, to facilitate system-wide, interdependent, and coordinated interventionsSuccessful implementation of organizational IPC strategies to support ASP and facilitate AMR mitigation and management, irrespective of resource setting, would require decisive organizational change management and integration of the ancient African philosophy of Ubuntu, meaning: ‘I am what I am because of who we all are’ :Firstly,Although the responsibilities of ASP and IPC programs are different, collaboration between these groups is essential to promote optimal outcomes. According to Manning et al, the vital work of IPC and ASP cannot be performed independently, and deliberate strategic relationship-building is essential . FurtherMultifaceted IPC interventions have been shown to be instrumental in reducing HOI rates, improving the rational use of antibiotics, increasing HH compliance, and reducing mortality in both developed and developing countries. Therefore, leveraging the butterfly effect of effective HH and IPC programs, can accelerate progress towards preventing the emergence and cross transmission of AMR bacteria, and can verify that an effective IPC program is fundamental to successful ASP in any organization.


Colorectal cancer is the cancer with the second highest and the third highest incidence rates for the female and the male, respectively. Colorectal polyps are potential prognostic indicators of colorectal cancer, and colonoscopy is the gold standard for the biopsy and the removal of colorectal polyps. In this scenario, one of the main concerns is to ensure the accuracy of lesion region identifications. However, the missing rate of polyps through manual observations in colonoscopy can reach 14%–30%. In this paper, we focus on the identifications of polyps in clinical colonoscopy images and propose a new N-shaped deep neural network (N-Net) structure to conduct the lesion region segmentations. The encoder-decoder framework is adopted in the N-Net structure and the DenseNet modules are implemented in the encoding path of the network. Moreover, we innovatively propose the strategy to design the generalized hybrid dilated convolution (GHDC), which enables flexible dilated rates and convolutional kernel sizes, to facilitate the transmission of the multi-scale information with the respective fields expanded. Based on the strategy of GHDC designing, we design four GHDC blocks to connect the encoding and the decoding paths. Through the experiments on two publicly available datasets on polyp segmentations of colonoscopy images: the Kvasir-SEG dataset and the CVC-ClinicDB dataset, the rationality and superiority of the proposed GHDC blocks and the proposed N-Net are verified. Through the comparative studies with the state-of-the-art methods, such as TransU-Net, DeepLabV3+ and CA-Net, we show that even with a small amount of network parameters, the N-Net outperforms with the Dice of 94.45%, the average symmetric surface distance (ASSD) of 0.38 pix and the mean intersection-over-union (mIoU) of 89.80% on the Kvasir-SEG dataset, and with the Dice of 97.03%, the ASSD of 0.16 pix and the mIoU of 94.35% on the CVC-ClinicDB dataset. Colorectal cancer is the cancer with the second and the third highest incidence rates for the female and the male, respectively. Early diagnosis has a huge impact on the survival from colorectal cancer . ColorecWith the rapid development of computer and information techniques, computer-aided diagnosis methods have been used in polyp segmentation tasks. However, the techniques of computer-aided polyp segmentations is still immature, especially in the cases that some complex and uncontrolled environmental factors exist. For example, existing computer-aided diagnosis methods can not effectively deal with factors that can affect the accuracy of polyp segmentation, such as intraluminal folds and variations of the polyp textures and locations. In , the priLater, some early machine learning methods were developed for colorectal polyp segmentations. However, these methods consumed a lot of computer memory and relied excessively on handcrafted features, thus were not robust enough. For example, the machine learning based methods in  got consIn the past few years, the deep learning technology has been adopted in the segmentations of medical images. Particularly, Olaf proposed the U-shape artificial neural network (U-Net) using an encoding-decoding structure . In the More recently, some network structures based on the U-Net were proposed for the medical image segmentations and showed outstanding performances . In partet al connected the encoding and decoding paths with dilated convolutions, where the receptive fields were expanded. However, the gridding effects could be introduced in this case. In order to solve this problem, P. Wang et al proposed the hybrid dilated convolution (HDC)  . The HDCon (HDC) . For exaon (HDC) . However1) We propose a novel N-shaped artificial neural network (N-Net) structure to conduct the lesion region segmentations of polyps in colonoscopy images. The proposed N-Net is designed based on the encoding-decoding framework. Within the proposed structure, the multi-scale information can flow between the encoding and decoding paths. The pretrained DenseNet modules based on the ImageNet are implemented in the encoding path of the N-Net to ensure the fast training convergence and good feature extraction performance of the entire network structure.2) To expand the receptive fields and facilitate the multi-scale information transmission between the encoding and decoding paths, we propose a strategy to design the generalized hybrid dilated convolution (GHDC). Compared with the existing works related to the dilated convolutions, the GHDC is established with a more flexible strategy to design cascaded dilated convolutional layers.3) Based on the strategy of GHDC designing, four GHDC blocks are designed to connect the encoding path and the decoding path. With experiments on two public available datasets: the Kvasir-SEG dataset and the CVC-ClinicDB dataset, we show that the GHDC blocks outperform the HDC in . MoreoveIn order to overcome the shortcomings of the current methods and optimize the colorectal polyp lesion region segmentation performance using colonoscopy images, we propose a novel N-shaped artificial neural network (N-Net) structure. It should be specially noted that although this work focuses on the colorectal segmentation task, the N-Net structure can be used generally for image segmentations. The N-Net structure is briefly described in In this paper, motivated by the U-Net encoder-decoder framework and the DenseNet modules, we propose a novel N-shaped architecture. As shown in In contrast to the U-Net structure, a “Conv (7 × 7)-BN-ReLU” operation  and an “Maxpool” operation (maximum pooling with the kernel size of 7 × 7 and the stride of 2) are implemented as the first stage of the encoding path. With the idea of transfer learning, the three encoding modules in the encoding path are designed based on the three dense blocks in the pre-trained DenseNet-121 from the ImageNet, respectively. Each dense block is composed by certain dense layers and a transition layer. The pre-trained dense layers in the three dense blocks and the first two transition layers are directly transferred. The number of dense layers in the three dense blocks are 6, 12 and 24, respectively. The number of output channels from the four encoders are 64, 256, 512 and 1,024, respectively. The feature map size from the Encoder 1 is 224 × 224 and is halved after the processing of each encoder.The decoding path in the proposed N-Net is composed with four decoding modules. Each of the decoding modules contains a “Conv (1 × 1)-BN-ReLU-TransposeConv (3 × 3)-BN-Conv (1 × 1)” procedure . The Decoder one contains two “Conv (3 × 3)” operations and a bilinear interpolation operation.i = 1, 2, 3, the outputs of the Encoder i and the GHDC block i + 1 are concatenated as the input of the GHDC block i. Besides, the outputs of the GHDC block i and the Decoder i + 1 are point-wise added as the input to the Decoder i.The encoding and decoding paths in the N-Net structure are connected with the proposed GHDC blocks, which are designed based on the novel strategy of GHDC designing in this paper. For With the connections between the encoding path and the decoding path using the GHDC blocks, the multi-scale features are deeply exploited with expanded respective fields. The detailed strategy of GHDC designing and the GHDC blocks are provided in the remainder of this section.In order to achieve sufficient multi-scale information transmissions, while expanding the respective fields without the gridding effects, we establish a more general and simplified strategy to design the cascaded dilated convolutions. In contrast to the HDC, flexible dilation rates and convolutional kernel sizes are enabled in the designed operation. Which is referred to as the generalized hybrid dilated convolution (GHDC) in this paper.k1, k2 and k3, respectively. The dilation rates used in the three layers are denoted as r1, r2 and r3, respectively.To explain the strategy of GHDC designing, we first provide a simple example to design the first three layers in a cascade of dilated convolutions in M2 and M3, respectively. According to M2 and k1 can be used to observe the coverage of the convolutional kernel in the first layer.In In the proposed strategy of GHDC designing, a key point is to ensure that all of the pixels in the feature map into the first layer are utilized in the operations in order to avoid the gridding effect.M3, k2 and r2 takes the value of 5, 3 and 2, respectively, then M2 becomes 2. In this scenario, by taking k1 = 3 and r1 = 1, it can be guaranteed that no holes exist during the cascaded convolutions.To meet this constraint, the parameters in the third and second layers are decided sequentially. For example, if lth (l > 0) layer in a dilated convolution cascade. In this scenario, it can be found that the sampling positions in this operation are influenced by the parameters of both the lth and (l + 1)th layers. Let us define Ml+1 as the maximum distance between neighbored sampled positions horizontally or vertically on the (l + 1)th (l > 0) layer, and define rl and kl to be the dilation rate and the convolutional kernel sizes on the lth layer, respectively. Then we can estimate Ml by the sliding of the dilated convolutional kernel on an xOy coordinate system. Due to the symmetry, there is no harm to simplify the problem by just observing the movement of the convolutional kernel on the x-axis.To be more general, we observe the dilated convolution operation on the arbitrary kl+1 pixels separated by the distance of Ml+1 on the feature map into the (l + 1)th layer, then the possible pixels covered by the convolutional kernel can be represented by: {}, where ith pixel covered by the convolutional kernel. In other words, if Ml+1, kl+1, rl, kl are determined, the parameter Ml can be derived.If we consider lth (l > 0) layer, we define Dl = {dl,i}, i = 1, … , klkl+1–1 to be a non-decreasing sequence. Then given Ml+1 and kl+1, rl and kl (l > 1) are determined with the following constraints:Ml, which is used to design the parameters of kl−1 and rl−1, can be determined as follows:Therefore, in the proposed GHDC model, the design of a dilated convolution cascade is done in a recursive manner. For the In addition, it is preferred that the receptive field in the first dilated convolution layer, which can be calculated using:For instance, if the kernel sizes of three are used in the cascaded dilated convolutions, we can get the following expression using i + 1 are consistent with that from the Encoder i , a GHDC transition layer, consisting of a bi-linear interpolation and a “Conv (3 × 3)-BN-ReLU” operation, is introduced.In the proposed N-Net, the GHDCs are conducted with four GHDC blocks between the encoding and decoding paths. As shown in ml,i and nl,i in In In order to verify the proposed N-Net, two publicly available datasets on polyp segmentations of colonoscopy images were used in the experiments: 1) the Kvasir-SEG dataset ; 2) the –4, 10–8, 16 and 300, respectively. The sum of the binary cross-entropy and the soft dice coefficient   to contrfficient  was usedn denotes the number of samples in a training data batch. yim and yim represent the predicted and actual probabilities that the ith pixel belongs to the class m , respectively.In To verify the performance of the proposed method, the metrics of Dice, ASSD and mIoU were implemented on the lesion region segmentation results in the experiments.S and G represent the region segmented by the N-Net and in the ground truth, respectively.The metric of Dice is defined as:Pa and Pb denote the set of boundary points segmented by the convolutional neural network (CNN) and in the ground truth, respectively. u to Pb.The metric of ASSD is defined as:TP, FP, TN and FN represent the numbers of pixels with true positive, false positive, true negative and false negative decisions for the lesion regions, respectively.The metric of mIoU is defined as:In order to verify the effectiveness of the N-Net structure and analyze the contributions of the GHDC blocks to the network performance, we first performed the ablation experiments with the GHDC blocks on the two datasets. The results of the experiments are shown in n = 4; the dilation rates and convolutional kernel sizes of the layers were taken as R1 = 1, R2 = 2, R3 = 5, R4 = 9 and k1 = k2 = k3 = k4 = 3, respectively). The experimental results are shown in Besides, we also conducted experiments to verify the advantage of the GHDC blocks compared the HDC on the two public colorectal polyp segmentation datasets. In particular, we compared the networks integrating the GHDC block 1 and/or the GHDC block 2 with the cases where the corresponding GHDC block(s) were/was replaced with the HDC(s). The HDCs in the experiments were implemented as proposed in   structure to conduct the lesion region segmentations of the colorectal polyps from colonoscopy images. To facilitate the multi-scale information transmissions, we propose a strategy of generalized hybrid dilated convolution (GHDC) designing which enables flexible dilation rates and convolutional kernel sizes to facilitate transmission of the multi-scale information. Based on the proposed strategy of GHDC designing, we design four GHDC blocks to connect the encoding path and the decoding path of the N-Net.The proposed method was evaluated on two publicly available colonoscopy image datasets for polyp segmentations: the Kvasir-SEG dataset and the CVC-ClinicDB dataset. The advantages of the proposed GHDC blocks were demonstrated through multiple sets of ablation experiments. In addition, the interpretability of the proposed GHDC blocks was analyzed through the visualization of the feature maps. Moreover, through comparative studies, the proposed N-Net was shown to outperform the state-of-the-art CNNs, including DeepLabV3+, TransU-Net and CA-Net, with the metrics of Dice, ASSD and mIoU as 94.45%, 0.38 pix and 89.80% on the Kvasir-SEG dataset and 97.03%, 0.16 pix and 94.35% on the CVC-ClinicDB dataset, respectively.In this paper, the research was conducted on two publicly available datasets of polyp segmentations for colonoscopy images, the Kvasir-SEG dataset and the CVC-ClinicDB dataset, where consistency were shown in the results. As the patient amount included in the two datasets was still limited, additional datasets  can alsoIn this work, we proposed an N-Net structure based on the encoding-decoding structure to conduct the polyp lesion region segmentations of colonoscopy images. In the proposed N-Net, the pre-trained DenseNet module was transferred as the encoding path of the network. In particular, we proposed a strategy of generalized hybrid dilated convolution (GHDC) designing to facilitate transmission of the multi-scale information and expand the respective fields. Based on the strategy of GHDC designing, four GHDC blocks were designed to connect the encoding path and the decoding path of the N-Net. Experiments were performed on two publicly available colorectal polyp lesion region segmentation dataset: the Kvasir-SEG dataset and the CVC-ClinicDB dataset. The advantages of the GHDC blocks were verified. Moreover, experimental results also showed that the proposed N-Net outperforms with a small amount of parameters, compared with the state-of-the-art methods.


Learning which environmental cues that predict danger is crucial for survival and accomplished through Pavlovian fear conditioning. In humans and rodents alike, fear conditioning is amygdala-dependent and rests on similar neurocircuitry. Rodent studies have implicated a causative role for dopamine in the amygdala during fear memory formation, but the role of dopamine in aversive learning in humans is unclear. Here, we show dopamine release in the amygdala and striatum during fear learning in humans. Using simultaneous positron emission tomography and functional magnetic resonance imaging, we demonstrate that the amount of dopamine release is linked to strength of conditioned fear responses and linearly coupled to learning-induced activity in the amygdala. Thus, like in rodents, formation of amygdala-dependent fear memories in humans seems to be facilitated by endogenous dopamine release, supporting an evolutionary conserved neurochemical mechanism for aversive memory formation. L-DOPA raclopride has been widely used to assess endogenous dopamine release during psychological tasks and pharmacological challenges. In contrast to previous genetic and pharmacological studies showing the involvement of dopamine in fear learning in humans, but unable to localize dopaminergic signaling to certain brain regions [11C]raclopride PET provides spatial resolution of dopamine release and allows pinpointing specific brain regions involved. Although dopamine release in the amygdala is scarcely investigated in humans, it should be noted that protein expression of the dopamine 2 receptor is high in the human basolateral nucleus [11C]raclopride BP has been reported to be high raclopride raclopride was injected through a venous catheter and followed by constant infusion during the 90 min of PET data acquisition. Following 50 min of resting PET data collection, participants underwent a differential fear conditioning paradigm during collection of blood-oxygenation-level dependent (BOLD) fMRI and skin conductance. The PET scanning continued 20 min after the fear conditioning paradigm.Participants were positioned supine in the combined Signa 3T PET/MR scanner  with their heads lightly fixated inside the head coil. A bolus (20 s) of the selective dopamine D2/3 receptor antagonist raclopride bolus-infusion protocol . Images were reconstructed into 18 5-min frames using ordered subset expectation maximization , including resolution recovery and a 5 mm Gaussian post-filter. Attenuation correction was done using the manufacturer’s atlas-based method, and all other corrections necessary for quantitative PET images were applied. Atlas-based attenuation correction has been shown to be less accurate than CT-based or zero echo time (ZTE) MRI-based attenuation correction, but because we were only addressing changes in receptor binding within the same patient and scan, this does not affect our results. Head movement in the scanner was restricted using foam cushions.Subjects underwent a 90 min PET scan on a 3 T Signa PET-MR scanner , initiated simultaneously with the start of a  and used as a measure of endogenous dopamine release. The analyses resulted in VOI values and parametric images of BPND, BP’ND and dopamine release. The parametric images were normalized to Montreal Neurological Institute (MNI) standard space in SPM12 by first co-registering each individual’s parametric images to their T1-weighted MRI image, then segmenting and normalizing the T1-weighted image to MNI space, and finally applying the transformation parameters to the PET images. Images were resliced to 4 mm isotropic voxels and smoothed with an 8 mm full width half maximum (FWHM) Gaussian kernel.PET data were analyzed both in VOIs and voxel-by-voxel using a nested two-step approach: first, the initial 50 min of the TACs were analyzed using the simplified reference tissue model (SRTM)  for VOIs11C]raclopride BPND), the time-activity data would follow the fit line of the 0–50 min data. As can be seen in Fig. ND. This deviation cannot be seen in the frontal cortex, thus not indicating dopamine release in this region.An illustration of the fit of the two SRTM models to mean TACs is presented in Fig. Ideally, the use of a bolus-infusion protocol results in a steady state, which would allow for use of simple radioactivity concentration ratios to measure occupancy instead of tracer kinetic modeling. However, the nested two-step SRTM method can account for deviations from steady state, which are nearly always present, whilst ratio methods cannot. Another, previously published analysis method (lp-ntPET)  would haND was assessed by performing a simulation study. For dopamine release levels resulting in a reduction in BPND of 0, 5, 10, 15 and 20%, 100 time-activity curves were numerically simulated using published values of rate constants, typical noise levels, and baseline BPND values of 2.6 (representing striatum), 0.3 , and 0.07 . Changes in BPND during the scan were simulated by a linear reduction in k3 between 50 and 70 min post injection, and resulting time-activity curves were analyzed using the nested version of SRTM. In addition, 100 TACs with a range of BPND changes between 0 and 30% were simulated for each of the baseline BPND values. A minor bias in post fear conditioning BPND of 0–2% and coefficient of variation (COV) of around 10% was found for baseline BPND values of 0.3 . For high baseline BPND (striatum), there was a positive bias in post-challenge BPND that was proportional to dopamine release levels, and a COV of around 2.5%. For lower baseline BPND , a varying bias of ±5% was seen with COV exceeding 40%. Hence, for striatum, the nested SRTM method seems to result in a proportional underestimation of dopamine release, whereas this effect is much smaller for amygdala .Robustness of measuring post-challenge BP1.Thus, simulations show a high accuracy and precision of the two-step nested method applied in the present work provided that the dopamine release time course, modeled as a gamma variate function, peaks before 70 min and has a relatively slow reduction after that. These conditions were confirmed both by microdialysis experiments in rodents  and by tFMRI data was preprocessed and modeled using SPM12 (fil.ion.ucl.ac.uk/spm/software/spm12) implemented in MATLAB R2018a . Each individual’s fMRI data was first slice-timing corrected, realigned and co-registered to the anatomical T1-weighted image, and then normalized to isotropic 4 mm voxels in the MNI standard space by applying the transformation parameters from the segmentation of the T1-weighted image. Finally, the images were smoothed with an 8 mm FWHM Gaussian filter.The first level model for each participant was fitted with onsets and durations of CS+, CS−, and shock (modeled as a stick function) convolved with the canonical haemodynamic response function from SPM12, together with the six realignment parameters from the realignment step, as regressors. Contrast images were created for CS+, CS− and US vs. baseline, and for the learning-related CS+ minus CS−. Mean BOLD responses within the regions of interest amygdala and striatum were extracted using the Wake Forest University School of Medicine (WFU) PickAtlas automatic anatomical labeling (AAL) definitions of these regions .11C]raclopride BPND, but could not detect any association in the amygdala  or in the striatum VOIs .To address potential influence of motion on the results, in addition to including the realignment parameters in the first level model, we set a cut-off to keep subjects with head motion not exceeding one acquisition voxel  in any direction . We alsoWe applied three a priori regions of interest in our analyses, the amygdala, striatum, and frontal cortex . The amygdala was our primary target based on our hypotheses, the striatum  was included based on the involvement in fear conditioning and rich dopamine receptor D2/3 expression in this region , and the11C]raclopride BPND VOI values and mean BOLD responses within the amygdala and striatum. We also applied voxel-wise paired t-tests (baseline vs post-conditioning) in SPM12 to locate overlapping voxels with both lower [11C]raclopride BPND  and learning-related neural activity (CS+ minus CS− BOLD responses) from fear conditioning. The statistical threshold was set to P < 0.05 family-wise error (FWE) corrected for multiple comparisons using random field theory and small volume correction within each region of interest .Our main analyses involved the raclopride PET we found decreased binding potential (BPND) from baseline to post conditioning in the amygdala  = 2.74, P = 0.007)  = 4.69, P = 0.0002)  = 0.54, P = 0.595)    Fig. , confirm02) Fig. , indicatF = 21.53, P = 0.0002) and Trial  = 23.96, P < 0.00001), and a CS× Trial interaction  = 2.25, P = 0.0023)  = 16.79, P < 0.001) and CS− = 13.77, P < 0.001), as well as an effect of CS for most trials, but importantly not for the first trial  (Supplementary Table r(16) = 0.44, P = 0.033, 95% CI: 0.05–1.00), consistent with a vast literature underscoring amygdala as a key brain structure for aversive memory formation in humans and other animals raclopride BPND with SCR difference scores (CS+-CS−) and found a positive linear relationship (r(16) = 0.60, P = 0.004, 95% CI: 0.27–1)  = 0.31, P = 0.209, 95% CI: −0.18 to 0.68). Percent change in amygdala [11C]raclopride BPND also predicted learning-induced neural activity in the amygdala (r(16) = 0.41, P = 0.044, 95% CI: 0.02–1)  = 0.29, P = 0.237, 95% CI: −0.20 to 0.67)  = 0.003, P = 0.991, 95% CI: −0.46 to 0.47)  = 0.27, P = 0.271, 95% CI: −0.22 to 0.66) , indicating specificity of dopaminergic facilitation of memory formation in the amygdala. No difference in dopamine-BOLD correlation coefficients between the amygdala and striatum could be detected .Next, we tested if dopamine release and learning strength are functionally coupled in the amygdala by correlating percent change in raclopride to measure dopamine release outside the striatum has been questioned. As mentioned in the introduction, this is based on the lack of change in BP in pharmacological occupancy studies. However, these studies have rarely included the amygdala or applied single scan bolus/infusion paradigms which arguably adds to the sensitivity of the measure. Moreover, test-retest reliability of amygdala [11C]raclopride BP has been reported to be high [11C]raclopride BPND in the amygdala and the striatum, although amygdala release had higher variability, whereas our control region, the frontal cortex did not indicate dopamine release. In addition, simulations  approach used in this study. A principal shortcoming of the nested SRTM method used here is that it is insensitive to transient changes in tracer binding, and thus only detects prolonged changes. In situations with transient changes, other methods such as lpnt-PET may be superior. Importantly, this does not affect the current results as the data showed prolonged reduction in tracer binding.Limitations of the current study deserve mentioning. First, the use of [ be high . Also, hL-DOPA administration strengthens safety memories during experimental fear extinction [Using simultaneous measures of dopamine release and neural activity in a combined PET/MRI scanner we show that dopamine release during fear conditioning is linked to aversive memory formation. This is in line with animal studies showing that dopamine is critical for fear learning  and suggtinction . Dopamintinction , instrumtinction , and in tinction . Thus, wSupplemental material