Helicobacter pylori (HP) in the oral cavity is discussed in some papers. The aim of study: To analyze the incidence of HPV and HP in oropharyngeal/laryngeal cancer persons versus persons with chronic tonsillar inflammation and healthy persons. Methods: The samples were taken in three groups: (1) tissue of oropharynx/larynx cancer (103 specimens); (2) tissue of palatine tonsils (85 specimens); and (3) healthy control group (50 specimens). We analyzed the presence of HP (PCR) and HPV genomic DNA  in the samples. Results: HP was detected in 86 samples (83.5%) and high-risk HPV in 62 samples (60.2%). We found a very high incidence of HP. In the cancer group, HP was detected in 82.5% cases and HPV positivity in 57.8%. In total, 7.2% of the cancer patients were negative for HP and HPV together. In turn, 53.6% of the cancer patients were positive for HP and HPV together. Four cases (4.2%) were positive for HPV only. VacA positivity was detected in 82 (79.6%) of the cancer cases and VacA negativity in 21 (20.4%) if the cancer cases. The incidence of HP in chronic inflammation (n = 85) was 65 cases (76.5%) and the incidence of HPV was 38 cases (44.7%). VacA positivity was detected in 59 (69.4%) of the chronic inflammation cases and VacA negativity was found in 26 (30.6%) of the chronic inflammation cases. Regarding the control group, we found HP positivity in 5 cases (11.1%) and HPV positivity in 19 cases (42.2%). There was VacA positivity in 6 cases (50.0%) of the control group. Statistically significantly lower prevalence of HP (p < 0.001) and HPV (p = 0.006) was found in the control group. Conclusions: We suggest that the palatine tonsils are colonized by HP. In our study, HP was present in oropharyngeal cancer in more cases in comparison with HPV infection. The presence of VacA from HP can have an influence on the human epithelial and immune cells’ regulation ways. Our results do not support idea that the CagA-positive HP is a primary carcinogen in oropharyngeal area.Objective: Oropharyngeal/laryngeal carcinoma are common cancers of the upper aerodigestive system. Human papillomavirus (HPV) is described as the most frequent in the cancer of unknown primary. The presence of  Oropharyngeal squamous cell carcinoma (SCC) and laryngeal SCC are some of the most common cancers of the upper aerodigestive system. Tobacco and alcohol consumption are the most important risk factors for both these cancers. The other risk factor for oropharyngeal cancer is human papilloma virus (HPV) infection, and this fact was the reason for modification of the tumor classification (TNM classification) and the treatment protocol of oropharyngeal cancer in the last decade. Other contributing factors may include viruses, bacteria, diet type, radiation exposure, gastro-esophageal reflux, occupation, and genetical inheritance . These iHelicobacter pylori (HP) in saliva [Helicobacter pylori can be a carcinogen or co-carcinogen of laryngeal SCC [Detection of n saliva  supportegeal SCC .Sivars et al. described that HPV-positive tonsillar and base-of-tongue SCC have a significantly better prognosis than HPV-negative patients . The mosAn HPV infection influences the presence of cell death molecules 8+ (CD8+) tumor-infiltrating lymphocyte counts and expression of HPV 16 was detected and used as a marker . The oraHelicobacter pylori was declared a type I carcinogen by the International Agency for Research on Cancer (IARC) in 1994 (IARC Working Group Reports). This bacterium was detected in the oropharynx, too [ynx, too ,8,9,10.Chen et al. described a malignant transformation-relevant role of HP in laryngeal precancerous lesions . On the Campylobacter-like organism (CLO) test, PCR genomic DNA detection, and reverse transcription polymerase chain reaction (RT-PCR), among others. The RT-PCR is reported to be the more exact method for detection of HP presence in tissue [HP was detected in pharyngeal and laryngeal pathologies ,13,14. Mn tissue ,13,14.The question about the local immunity depression and/or any other influence on the immunity and cell biology is real. HPV is a carcinogen in the oropharyngeal area. This fact was described in many papers ,18,19,20(1)To analyze of incidence of infection by HPV and HP in oropharyngeal and laryngeal cancer patients. Can we compare the incidence for cancer patients and chronic tonsillar inflammation to healthy persons?(2)Helicobacter pylori presents in oropharyngeal and laryngeal cancer patients. Determining the incidence of cytotoxin-associated protein (CagA), vacuolating cytotoxin (VacA), and the HPV subtypes in the cancer cohort and chronic inflammation group compared to healthy persons.Analysis of the HPV serotypes present in oropharyngeal and laryngeal cancer patients. Genomic analysis of The aim of the study:This study has a similar methodology as our previous manuscripts based on this grant ,21,22,23Helicobacter pylori and serotype of HPV detected in oropharyngeal and laryngeal cancer and chronic inflammation patients” was conducted with the consent of the Ethics Committee of the Motol University Hospital, Czech Republic—project number IGA NT-11523/6-2010.The authors declare that the prospective study “Genome of Helicobacter pylori and serotype of HPV detected in oropharyngeal and laryngeal cancer and chronic inflammation patients” took place with the consent of the patient. Each patient provided informed consent for accordance into the study. All of them are in accordance with the Helsinki protocols. The authors declare that the prospective study “Genome of Inclusion parameters ,21,22,23Patients with oropharyngeal SCC. The patients were included across all nodal statuses (N0–N3) but no presence of distant metastases. Patients with chronic tonsillar inflammation and/or patients with tonsillar infection recurrence were observed minimally two times yearly in the last five years or three or more times in the last two years. No antibiotic therapy was given in the six months preceding surgery. The tissue samples were taken after removing the primary tumor by a surgical procedure in cancer patients, and in the chronic inflammation group were taken after tonsillectomy from chronic tonsillitis patients. The saliva samples (control group only) were taken from the healthy persons group without any chronic inflammation in their patient history. No signs of cancer were present in the patient’s history ,21,22,23Child and adolescent patients were excluded (age under 18).Pregnant women were excluded.The patients with any of the following: human immunodeficiency virus (HIV), Hepatitis B or C viruses.Salmonella infections, tuberculosis, or zoonosis were excluded.Any chronic The patients of any other cancer in history were excluded.The patients with any treatment of the immune system, such as HIV , and patients on immunosuppression therapy (incl. steroids) were excluded.The patients suffering from chronic gastric inflammation and/or ulcers. The persons with a history of from reflux  were excluded.Antibiotic treatment administered within six months preceding the surgery or sample collecting.Exclusion’s parameters used for all groups:(a)Tissue of cancer by patients undergoing surgical treatment of oropharyngeal SCC.(b)Tissue of tonsils by patients undergoing surgical treatment of chronic tonsillitis, suffering one infection yearly for more than 5 years, or 2 infections in the last 3–5 years, or 4–5 infections in the last two years.(c)For the control group, we used the saliva samples from patients coming for a routine checkup appointment.Clinical selection of samples was done in three groups: Samples were taken from patients suffering from oropharyngeal SCC and chronic tonsillitis (TCR). For the control group, saliva samples from patients with non-oropharyngeal diagnoses were collected ,21,22,23The samples were stored in the Department of Otorhinolaryngology and Maxillofacial Surgery, 3rd Faculty of Medicine Charles University and Military University Hospital Prague. All surgical teams were instructed to handle tissue biopsies according to identical protocols ,21,22,23Collection of tissue specimens was done using sterile instruments at the start of surgery prior to application of any drug disinfection substances into the oral cavity ,21,22,23R M4RT Collection and Transport Medium. These specimens were transported into the laboratory where they were vortexed, aliquoted, and the nucleic acids isolated using a Roche MagNAPure Compact  automated isolator. The following procedure involved the real-time PCR technique for Helicobacter pylori detection and genotyping [For collection of samples, we used Remel Microtestnotyping ,21,22,23Helicobacter pylori flagellar gene by commercial real-time PCR and commercial real-time PCR screening tests for HPV, based on fla gene detection [We developed and optimized the tests and procedures in co-operation with TIB MolBiol Berlin, Germany. Samples and nucleic acid isolates were stored at −80 °C ,21,22,23etection ,21,22,23vacA gene was genotyped from the middle and final regions according to the technology described in the literature [With regard to genotyping, the terature ,21,22,23The primers used for PCR detection of HP were used according to van Doorn et al. . For detHelicobacter pylori Fla-Format Assay  was used. Helicobacter pylori real-time PCR assay probes were used: cagA, vacAs1a, vacAs1b, vacAs2, vacAm1, and vacAm2, comprehensively described in published papers [The commercial real-time PCR assay BIORON Real-Line d papers ,21,22,23Detection was done by Sacace HPV High-Risk Screen Real-TM Quant Assay . Assays were performed according to technology instruction and according to published technology ,21,22,23Helicobacter pylori assays primer sequences was used for cag AF+, cagAR−, VAF1F+, VAXR−, HPMGF, and HPMGR. This has been comprehensively described in the literature [terature ,21,22,23p-values < 0.05 were considered statistically significant in all statistical analyses.All statistical analyses were performed using IBM SPSS statistics . The data were analyzed using descriptive methods, Fisher’s exact test (two-tailed), and Pearson’s chi-square test. Helicobacter pylori DNA sequences . On the other hand, we analyzed the DNA of the high-risk types of HPV groups A9, A7, and A5/6. The incidence of both HP and HPV is summarized in This study analyzed 103 samples taken from squamous cell carcinoma in oropharyngeal area, 85 specimens taken from chronic inflammation of palatine tonsils, and 50 specimens of saliva from healthy persons (control group specimens), using techniques for detection of p < 0.001) and HPV (p = 0.006) in the control group.These data show evidence of HP presence in the cancer tissue in oropharyngeal area, especially in the lymphoid one. We can conclude that the significance of a higher rate of HP presence (83.5%) is based on improved laboratory techniques and methods of sampling, and on the other hand based on a higher incidence of this infection in tissue generally. A higher presence as also observed in chronic tonsillar inflammation patients (76.5%). This incidence is significantly higher compared to the control group (HP incidence 12.0% only). Statistical result: statistically significantly lower prevalence of HP  are displayed in On the other hand, VacA S1a,bM1 presents in 41.9% of CagA-negative cases compared to VacAS1,bM2 that presents in 24.4%. This result suggests that the presence of CagA-negative VacA genotypes was higher than the CagA-positive and VacA genotypes generally. This may reinforce the hypothesis that VacA positivity of HP can be the reason for the pathogenicity of HP in oropharyngeal malignancies.Helicobacter pylori genotypes in the chronic inflammation cases (Group B) is presented in The confirmation of VacA pathogenicity detected by analysis of VacA gene of Helicobacter pylori can play a role in the pathogenicity of Helicobacter pylori in oropharyngeal malignancies.Incidence of the VacA S1bM1 genotype is the most often and presents in 27.7% of cases in Group B, independently of the CagA genotype. On the other hand, in CagA-negative cases, the presentation of VacAS1a,bM1 is in 30.8% of cases compared toVacAS1bM2 in 33.9%. These results suggest that the CagA-negative VacA genotype cases were present in the same incidence see . This faThe presence of different genotypes of HP in Group A and Group B compared to the control group suggest the association of HP infection with moderate immunological status of the lymphatic tissue and/or the local immunity see . The canHelicobacter pylori was detected in 86 samples (83.5%), while high-risk HPV in 62 samples (60.2%). The HPV positivity A5/6 and any HP positivity together were detected in 38.8% of cases in Group A. HPV positivity A7 was detected in 16 cases (15.5%) and HPV positivity A9 in 12 cases (11.6%). Helicobacter pylori was genotyped as a sole agent in 34 samples (32.98%). HPV A5/6, a7, and A9 was detected as a sole agent in 7 cases (6.8%) only. A high number of papers reported HPV type 16 as the main cancer inductor. HPV type 16 is included the A9 group. In the used tests, we used a specific fluorescent signal for detecting the A9 group. We detected HPV type 19 in the A9 group only in 13 cases out of 60 HPV-positive cases. The A5/A group was detected in 46 cases (44.6%) out of 60 HPV-positive cases. The A7 group was detected in 21 cases (20.4%) out of 60 HPV positive cases.DNA of Chen et al. described the microbiota role in four cancer types  . This BaLopez et al. detected the circulating markers after resection of HNSCC . The anaThe incidence of HP in the tonsil tissue in the chronic inflammation group is of a lower rate than cancer patients.Helicobacter pylori.Payäo et Rasmussen described extra gastric reservoirs of HP in saliva and dental plaque, the tongue, and oropharynx . The autHelicobacter pylori-negative samples. HP was detected in 77.6% of patients in this group. HPV was positive in 47.1% samples. On the other hand, Hwang et al. published a meta-analysis with no finding of HP colonization in tonsillar tissue [The presence of HP in the oropharyngeal area, chronic tonsillitis, and oropharyngeal tumors has been described by us in many publications ,10,21,22r tissue . The metr tissue . Siupsinr tissue . The conr tissue .Helicobacter pylori in tonsillar tissue samples of patients with chronic tonsillitis is controversial.The role of HP infection in adenoid and tonsillar tissue is unclear yet. The incidence of HP in tonsils is not strong evidence of the influence of HP to immune cells in the oropharynx. On the other hand, the presence of CagA-positive HP in stomach area associates with a cancerogenic effect. This fact can relate to some influence on the local immunity in the tonsils and/or oropharyngeal area. The sharp difference in HP presence in the oropharyngeal area published in many papers depends on methods of HP detection. The data in our study support that the “gold standard” for oropharyngeal area is the PCR detection of the HP. Naserpour Farivar et al. described the colonization of tonsils in chronic tonsillitis patients by HP . These aHelicobacter pylori and HPV. To the contrary, 53.6% of cancer patients were positive both for HP and HPV. Only four cases (4.2%) were positive for HPV only.The results of this study showed a very high incidence of HP detected by PCR resp. rt-PCR. In the cancer group, the HP was detected in 82.5% cases. In the same group there was HPV positivity in 57.8%. Only 7.2% of cancer patients were negative both for If we compare these results to the control group of saliva analyses of healthy persons, we can see that HP positivity was detected in 5 cases (11.1%) and HPV positivity in 19 cases (42.2%). A saliva sample was used for the control group because no tissue samples could be taken in healthy patients. The HP detection in saliva was described in the literature . The useWe suggest that the fact about the incidence of HP in the oropharyngeal area is described in the present study.vacA and cagA genes. Our analysis focused on the incidence of the CagA and VacA cytotoxins. The presence of CagA-positive HP was in 23 cases (23.7%). CagA was described as the main carcinogenic factor in the stomach. Hatakeyama described the main carcinogenic process in stomach epithelial cells by bacteria through the effect of tyrosine phosphorylation at the Glu-Pro-Ile-Tyr-Ala (EPIYA) [cagA gene-encoded CagA protein is delivered into the gastric epithelial cells. The CagA intracellular effect promotes the neoplastic transformation in epithelial cells in the stomach. The role VacA in the epithelial human cells has not been described yet. In the literature described, some have studied an association between the vacA s1/m1 strains and gastric adenocarcinoma [We have analyzed the HP genome, respectively, the  (EPIYA) . This waarcinoma ,36. The arcinoma . The Vacarcinoma . The actarcinoma .Helicobacter pylori than HPV lead to more questions. We cannot asign HP to be a carcinogen for the oropharyngeal area yet. The evidence requires further studies. The cancerogenic effect of HPV has been declared in the literature, with strong evidence.The significant difference among the presence of HP in the respective group and the higher incidence of Considering our data, we suggest the revision of the theory on HPV-induced oropharyngeal cancer. Our results should support the hypothesis about the influence of HP (VacA) on local immunity. It seems evident that HP is a long-term resident in the oropharyngeal area and tonsils. We suggest that HP should be seen as a contributor who could switch on the process, resulting in cancerogenesis.We believe that the results in this study support the following conclusions:Helicobacter pylori. The high frequency of detected DNA of HP in the tissue of cancers and tonsillar tissue is the reason for these conclusions that HP is a resident in the oropharyngeal area. We suggest that the tonsils  can be colonized by The presence of HP in oropharyngeal cancer is real in more cases compared to HPV infection.The majority of HPV-infected patients has an HP infection at the same time in the same sample.The presence of HPV in oropharyngeal cancer patients alone is rare.Data in the study suggest that HP can influence local immune cells and may cause a decrease in immunity towards HPV and/or any other infectious agents.The CagA-positive HP was detected in less cases. This result of the study does not support the idea that CagA-positive HP is a primary carcinogen in the oropharyngeal area.The high number of HP patients in the oropharyngeal cancer group compared to the HPV infection must be explained in the next study.The incidence of HP infection and the presence of different genotypes suggest the association of infectious agents with oropharyngeal cancer.


Epidemiologic studies in Puerto Rico have documented the prevalence of human papillomavirus (HPV) infection on the island, reporting rates up to 79% . RegardiParallel with these scientific efforts to estimate the impact of HPV in Puerto Rico, grassroots movements have promoted educational campaigns and outreach activities to increase HPV vaccine uptake and awareness at the community level, as well as developed training opportunities among health care professionals. These efforts from multiple sectors  led Puerto Rico to have among the highest HPV initiation rates since 2014. These sectors have been pioneers in developing policies that support HPV vaccination access in Puerto Rico. Among the more meaningful is Law No. 9 , which provided HPV vaccine access to girls aged 11 to 18 years. The law was amended in 2012 to also include boys aged 11 to 18 years .In Puerto Rico, under Law No. 25 , the Puerto Rico Secretary of Health can decide which vaccines will be required for school entrance . The lawNotably, there was a 1-year gap between the public notification and the policy adoption. Given this time frame, several groups voiced their concerns against the mandate  and publicly expressed their apprehension about the new requirement. Research has shown that this period permitted the development of bills intended to deter or “water down” the new requirement   to accomplish. Moreover, the death of a young mother of 3 from cervical cancer in 2015 catalyzed a movement led by VOCES to advance the work surrounding HPV vaccination . AnotherOur team conducted key informant interviews with implementers . We also interviewed participants who were opposed to the policy to explore the factors that facilitated or hindered the successful implementation of the HPV vaccine school-entry requirement in Puerto Rico. The principal barriers identified were misinformation about the HPV vaccine and lack of knowledge about the implementation process of the mandate. In contrast, people’s principal concern against the policy was the government’s excessive interference in deciding what was best for their children. During this activity, the key informants’ recommendations were to 1) inform parents about the pros and cons of the vaccine; 2) actively use social media as a channel for dissemination of reliable information, acknowledging how this media is also used to spread misinformation; 3) train school staff; and 4) identify a central-figure school staff member who can follow up on the completion of doses and guide the parents. Additional findings indicated parental hesitancy, lack of awareness about the HPV vaccine and the new school-entry vaccination requirement, and unfounded concerns about the potential side effects of the HPV vaccine .Preliminary analyses using data from the Puerto Rico Immunization Registry (PRIR) showed an increase of 54.0% in vaccinations among children aged 11 to 12 years when comparing the pre-policy (2017) and post-policy 2019) periods. Nevertheless, several challenges are worth discussing. First, HPV vaccine implementation was 11 months after Hurricane Maria, one of the worst natural disasters in Puerto Rican history. The impact of this hurricane on the school and public health systems led to an estimated 2.43% population migration to the continental US in 2017  . This mi periods.The recent implementation of policy that establishes an HPV vaccine requirement for school entry in Puerto Rico provides a unique opportunity to assess the impact on HPV vaccination rates on the island and to explore the broader context of HPV vaccine school-entry requirements in other US states and territories. This implementation also encourages the strengthening of partnerships between local communities and policy makers to create additional strategies to support this requirement and address challenges. Educational programs about HPV vaccination must be disseminated to reduce public skepticism and clarify misinformation among hesitant parents. Furthermore, epidemiologic studies should be developed to assess the impact of changes in HPV vaccination rates on future incidence of HPV-related precancers and cancers in Puerto Rico, with the potential to document the progress toward eliminating cervical cancer on the island.


Diabetes is linked with poorer cervical cancer prognosis, and people residing in the Southern region of the U.S. are disproportionately diagnosed with diabetes and cancer. The HPV test was recently recognized as the preferred method of cervical cancer screening by the American Cancer Society. Through our observational study, we sought to investigate the HPV testing behaviors among females with and without diabetes across the U.S. Our nationally representative estimates reveal that less than half of females reported HPV testing, and females with diabetes in the Deep South have the lowest rates of HPV testing. Various risk factors were identified to significantly lower the odds of HPV testing, including a diabetes diagnosis, older age, living in the Southern region of the U.S., and absence of certain comorbidities. The lower rates of HPV testing among females with diabetes, especially those living in the Deep South, leave these populations vulnerable to cervical cancer.p < 0.001). Screening rates with HPV tests were lowest among females with diabetes in the South in 2016 (29.32% (95% CI: 26.82–31.83)), 2018 (39.63% (95% CI: 36.30–42.96)), and 2020 (41.02% (95% CI: 37.60–44.45)). Conclusions: Females with diabetes are screening with HPV tests less frequently than females without diabetes, and females living in the South, particularly states in the Deep South, report the lowest rates of HPV testing.Background: Due to diabetes being linked with poorer cervical cancer prognosis, this study aimed to evaluate HPV testing behaviors among females with and without diabetes across the U.S. by geographic area in 2016, 2018, and 2020. Methods: This cross-sectional study used the Behavioral Risk Factor Surveillance System (BRFSS) from 2016, 2018, and 2020. The study population included females aged 25–69 years old, stratified by self-reported diabetes status. The primary outcome measure was cervical cancer screening behavior, which was evaluated by self-reported HPV test uptake/receipt (yes/no). Results: A total of 361,546 females from across the U.S. were sampled. Within the study population combined from all study years, the overall likelihood of receiving an HPV test was significantly lower among females with diabetes [37.95% (95% CI: 36.87–39.04)] compared to those without diabetes [46.21% (95% CI: 45.84–46.58)] ( Diabetes continues to be a significant health challenge in the U.S., where the prevalence has consistently increased over time . Some prThe proposed relationship between diabetes and cancer has drawn experts’ interest for a long time, as the two are frequently co-morbid conditions ,8. AlthoCervical cancer is burdensome to females across the U.S. A total of 12,733 females were newly diagnosed with cervical cancer in 2018 alone, and the cervical cancer mortality was 4138 . In 2021While the burden of cervical cancer remains alarming, the number of cervical cancer cases has declined since the introduction of the HPV vaccine in 2006 ,20,21 anCancer screening serves as an effective tool for early cancer detection, and uptake of cervical cancer screening practices can have public health implications on cancer incidence and mortality outcomes . The twoHPV is now recognized as the cause for most cervical cancer diagnoses , so testParticipation in proper cervical cancer screenings among persons with diabetes could allow for the best chance at early detection and a more favorable prognosis. Despite this, there have been limited studies published that address these behaviors in this population. Through systematic review and meta-analysis, one study found that females with diabetes were significantly less likely to screen for cervical cancer than females without diabetes . HoweverOur study aims to evaluate cervical cancer screening behaviors among females with and without diabetes across the U.S. by investigating their HPV testing practices in 2016, 2018, and 2020. Due to the disproportionately high diabetes rates and low utilization of cervical cancer screening practices in various regions of the U.S. ,34, estiThis cross-sectional study used the Behavioral Risk Factor Surveillance System (BRFSS) data to evaluate cervical cancer screening behaviors among females in the U.S. The BRFSS is a cross-sectional telephone survey administered by the CDC that provides standardized questions regarding risk behaviors and preventive healthcare practices among a nationally representative sample . AdditioThe inclusion criteria remained broad to maximize external validity, so the study population included adult females aged 25–69. This study population was chosen by following the ACS guidelines for primary HPV testing to include survey respondents that were the correct biological sex for the respective cancer screening and within the age range of 25–65 years old , which rThe primary outcome measure was cervical cancer screening behavior, which was evaluated by self-reported HPV test uptake/receipt. The question used was, “An HPV test is sometimes given with the Pap test for cervical cancer screening. Have you ever had an HPV test?” The outcome was categorized as yes for those who responded “yes” to HPV test screening, no for those who responded “no” or “don’t know/not sure,” and missing for those who “refused” or did not answer the question.n) to represent the actual number of BRFSS respondents and weighted (weighted n) to represent the population after considering the BRFSS sampling design  than females without diabetes  (p < 0.001).In 2016, 2018, and 2020 BRFSS samples, 361,546 females aged 25–69 years old met inclusion criteria for the study population. A total of 41,442 females self-reported diabetes, and the remaining 320,104 self-reported not having diabetes . The disp < 0.001). Looking at each year separately, p < 0.001 for all). Across the entire study population, screening rates with HPV tests were lowest among females with diabetes in the South in 2016 (29.32% (95% CI: 26.82–31.83)), 2018 (39.63% (95% CI: 36.30–42.96)), and 2020 (41.02% (95% CI: 37.60–44.45)).Within the study population combined from all study years, the overall likelihood of receiving an HPV test was significantly lower among females with diabetes [37.95% (95% CI: 36.87–39.04)] compared to those without diabetes [46.21% (95% CI: 45.84–46.58)] . In 2018, females with diabetes had significantly lower rates of HPV screening compared to those without diabetes in Georgia, Louisiana, and South Carolina . In 2020, females with diabetes had significantly lower rates of HPV screening compared to those without diabetes in Alabama, Louisiana, and Tennessee .To better understand current cervical cancer screening behaviors across the U.S., this study investigated HPV testing practices among females with and without diabetes. Overall, HPV testing rates did not meet the target cervical cancer screening rate of 84.3% proposed by Healthy People 2030 . The HeaThe strength and significance of this study are found in the timely investigation of cervical cancer screening with the HPV test based on recent guideline updates from the ACS in 2020, along with the nationally representative nature of the findings. The novelty of this study lies in our investigation of HPV testing behaviors between females with and without diabetes. We found diabetes to be significantly associated with a lower likelihood of HPV testing, which is similar to previous studies that found strong associations among females with diabetes and lower cervical cancer screening rates ,43. An iWhen comparing regional HPV testing rates to national rates, the South exhibited the lowest screening rates in all years. Various states in the Deep South demonstrated significantly lower screening rates for females with diabetes versus females without diabetes, such as Alabama, Georgia, Louisiana, South Carolina, and Tennessee. These differences in screening rates by diabetes diagnosis changed across years in most states in the Deep South. Notably, the marked difference persisted across all years in Louisiana. Overall, the states in the Deep South make up a majority of the “diabetes belt” with the highest prevalence of diabetes in the U.S. . TherefoAdditionally, several points arise when discussing the impact of other factors on females performing appropriate HPV tests. We found Asian females to have lower odds of testing than Whites, and a previous study also reported lower cervical cancer screening rates among Asian Americans . We founDespite the concerns mentioned, it is also important to note that overall HPV testing rates have increased over time. This finding is similar to previous evaluations of co-testing, where rates of HPV test in this method increased after a change in ACS screening guidelines for cervical cancer in 2012 . From 20While we closely adhered to ACS guidelines, we could not make hard cutoffs for their age recommendations. All females within the 65–69 age category were included to capture females 65 years old. This led to the over-inclusion of females outside of the recommended ages in the ACS screening guidelines. With the self-reported responses in BRFSS, inaccurate classifications of variables, including diabetes diagnosis, could be present. Despite this concern, surveys with self-reported healthcare data, such as BRFSS, have still been found to have high reliability, particularly in preventive testing and diagnoses of chronic diseases/conditions . It mighFemales with diabetes are screening for cervical cancer with the HPV test less frequently than females without diabetes. Further, females living in the South reported the lowest rates of HPV testing, particularly in states in the Deep South. These populations of females with diabetes remain vulnerable to cervical cancer, so preventive measures must be taken through proper cervical cancer screenings. Future projections of diabetes prevalence, age, and regional disparities create a need for healthcare providers to adhere to the recent ACS guidelines that favor the primary HPV test. With the new recommendations, there may be a continued increase in HPV screening among persons with and without diabetes. However, critical work must be done to reach the currently projected goal for cervical cancer screenings of 84.3% from Healthy People 2030 , and spe


To determine the impact of Human Papilloma Virus (HPV) oropharyngeal cancer (OPC) status on the prediction of head and neck squamous cell cancer (HNSCC) chemoradiotherapy (CRT) outcomes with pre-treatment quantitative diffusion-weighted magnetic resonance imaging (DW-MRI).n = 57) and largest pathological node (n = 59) were placed to analyse ADCmean and ADCmin. Unpaired t-test or Mann–Whitney test evaluated the impact of HPV OPC status and clinical parameters on their prediction of post-CRT 2 year locoregional and disease-free survival (LRFS and DFS). Multivariate logistic regression compared significant variables with 2 year outcomes.Following ethical approval, 65 participants  underwent pre-treatment DW-MRI in this prospective cohort observational study. There were 46 HPV OPC and 19 other HNSCC cases with Stage III/IV HNSCC. Regions of interest (ROIs)  at the primary tumour (mean was predictive of 2 year LRFS (p = 0.04). However, only the HPV OPC diagnosis  predicted outcomes on multivariate analysis. None of the pre-treatment ADC values were predictive of 2 year DFS in the HPV OPC subgroup (p = 0.21–0.68). Amongst participants without 2 year disease-free survival, HPV-OPC was found to have much lower primary tumour ADCmean values than other HNSCC.On univariate analysis of all participants, the primary tumour area ADCKnowledge of HPV OPC status is required in order to determine the impact of the pre-treatment ADC values on post-CRT outcomes in HNSCC.mean and ADCmin values acquired using different ROI methods are not predictive of 2 year survival outcomes in HPV OPC.Pre-treatment ADC However, treatment fails in more than 30% of advanced stage tumours, and salvage surgery for locoregional recurrence has a potentially poor outcome.2 Identifying tumours that may not respond to CRT provides prognostic information, guides more proactive surveillance, and prompts more intensive or modified treatment plans such as radiotherapy dose escalation or surgery.Head and neck squamous cell cancer (HNSCC) is the seventh commonest cancer worldwide.mean in predicting HNSCC treatment response.3–25 Whilst many have demonstrated that that a high pre-treatment ADCmean predicts a poor outcome following CRT,6–8,11,12,15,18–21 there are conflicting results.3–5,9,10,13,14,16,17,22–25One potential imaging biomarker is the pre-treatment tumour apparent diffusion coefficient (ADC) on diffusion-weighted MRI (DW-MRI). A number of studies have assessed the value of pre-treatment ADC26 may both influence ADCmean measures27–30 and clinical outcomes. HPV OPC is increasing in incidence and now accounts for 70–80% of OPC in the United States and Western Europe.31 Whilst the HPV OPC status is rarely documented in studies of pre-treatment ADC values in predicting HNSCC outcomes, recent evidence has suggested it to be a potentially important factor.4,14,15,24Human Papilloma Virus (HPV) oropharyngeal cancer (OPC) status may be an important confounding factor in these studies, since its unique histopathological characteristicsOur primary objective was to determine the impact of HPV OPC status on the ability of pre-treatment DW-MRI ADC values to predict 2 year survival outcomes in Stage 3 and 4 HNSCC following CRT.http://www.controlled-trials.com/ISRCTN58327080) following Research Ethics Committee approval (REC reference 13/LO/1876) and informed consent.Participants were recruited for a prospective single centre cohort observational study between 2014 and 2017 . Exclusion criteria were prior chemoradiotherapy, an ECOG performance status >2, lack of capacity to understand the patient information sheet or inability to provide informed consent, and known allergy to gadolinium-based contrast medium or eGFR <30 ml/min.Participants were eligible if there was histologically confirmed Stage 3 or 4 primary HNSCC without distant metastases, an axial section delineating an 1 cm5,7,11,12,19 the ADC measurements were predicted to have a standard deviation of 40 and to demonstrate a difference of 50 × 10−6 mm2/s between participants with and without 2 year survival outcomes. Using a 5% significance level and 80% power, it was calculated that at least 40 subjects would be required for the study without any accounting for loss of data.Sample size calculation was based on 70% of patients demonstrating 2 year disease free survival (DFS). Based on previous reports,32 HPV status was evaluated with p16 testing using an immune-stain or high-risk HPV DNA testing using in situ hybridisation. Biopsies at diagnosis were obtained from the primary tumour (n = 56), lymph node (n = 7) or both (n = 2). Intensity modulated radiotherapy (IMRT) was delivered as per the standard of care which was 7 Gy in 35 fractions; 2 Gy per fraction delivered once daily, 5 days a week. Concomitant intravenous cisplatin at a dose of 35 mg/m2 every 7 days, starting on Day 1 of radiotherapy, was administered in all patients with adequate GFR and no contraindications (n = 47) with carboplatin (n = 16) or radiotherapy alone (n = 2) being used in the remaining participants.HPV status was analysed for all OPC as per standard of care. Non-OPC HNSCC was not routinely tested for HPV status, according to international guidelines.4,13 This approach was adopted, since HPV OPC is a distinct entity with unique histopathology and favourable treatment outcomes relative to all other HNSCC and the prognostic impact of p16 positive status is restricted to OPC only.33,34The cohort was subsequently divided into HPV OPC and all other HNSCC (including HPV negative OPC) for analysis as in previous studies.2) and the following scan parameters: repetition time 5900 ms, echo time 60 ms, two signal averages, FOV 240 x 240 mm, slice thickness 4 mm with a 0.5 mm slice gap. Mono-exponential ADC maps were calculated from the b = 100 and b = 800 values.Participants underwent 1.5 Tesla MRI  using a surface phased array 20 channel neck coil. The routine MRI protocol  was suppThe ROIs were placed by a radiologist  under the supervision of another radiologist . The first 22 primary tumours and 24 nodes were also independently analysed by another radiologist  to assess interobserver agreement.b = 800 s/mm2 map, but with access to the post-gadolinium fat saturated T1 axial sequence. Three separate ROIs were placed individually within the primary tumour and/or largest pathologically appearing lymph node (Volumetric ROI (vROI) placed on multiple sections to encompass the whole lesion volume.Area ROI (aROI) placed around the largest single cross-sectional area of the lesion.b = 800 s/mm2 map but excluded any areas of necrosis defined by cross-referencing to areas of either high signal on the b = 0 map or absence of gadolinium enhancement.Representative area ROI (rROI) placed on the largest cross-sectional area in the core of the lesion. This focused on an area of increased DWI signal on the Free hand ROIs were placed using OsiriX v. 8.0.2, open source Mac-based medical image processing software with the images magnified to a standard 300%. They were defined on the DWI mph node :Volumetrb = 0 map and avid gadolinium enhancement were avoided on all ROIs. An ADC map was generated from the b100 and b800 s/mm2 images to mitigate the perfusion effects. ADCmean and standard deviation were recorded with ADCmin calculated as ADCmean – one standard deviation (SD).21 The vROI was also used to define the volume of the primary tumour and/or largest lymph node using the summation of areas technique. A ROI was also placed within the cervical spinal cord on the ADC map, and ADCmean was calculated as a reference for any non-patient related variability or artefact.Areas of peritumoural inflammation characterised by high signal on the Outcome evaluation comprised clinical assessment at 1 year and 2 years following completion of treatment. Treatment failure was determined by cytological or histological confirmation (biopsy or resection) or by serial progression on imaging follow-up. The 2 year locoregional recurrence free survival (LRFS) and 2 year disease free survival (DFS) were recorded according to the status at 2 years following completion of treatment.18F-FDG PET-CT was also obtained. This comprised a 6 mm diameter volume of interest (VOI) being placed at the site of most intense FDG uptake within either the primary lesion and/or the largest lymph node. If there was reduced uptake on the post-treatment images relative to background, a 6 mm VOI was placed at the same site as the post-treatment MRI ROI. If necrosis was identified within a lesion, the area of necrosis was excluded. The SUVmax was calculated with semi-automated software on a Hermes workstation .A standard of care 12 week post-treatment positron emission tomography computed tomography (PET CT) study was used to guide clinical management. A quantitative evaluation of the post-treatment p value of < 0.05 being considered statistically significant.Statistical analysis was performed using Stata (v. 15.1) with a Descriptive statistics summarised the patient demographics, HPV status, tumour site and staging.mean and ADCmin values obtained using the different ROI methods at both primary tumour and nodal locations were analysed for all participants, and for HPV OPC and other HNSCC subgroups. ADC values were compared between HPV OPC and other HNSCC participants.ADCp < 0.2) were used as independent variables in the multivariate analysis.ADC values, HPV OPC status and clinical variables  were compared with the dichotomised 2 year outcomes of DFS and LRFS. Any variables associated with 2 year survival outcomes  for each of the vROI, aROI and rROI at primary tumour and nodal locations were evaluated in the sample analysed by two observers.The participant flowchart is summarised in There were 101 patients eligible according to study criteria with 70 patients enrolled, but five were subsequently withdrawn. Of the 65 participants , there were 54 (83%) Stage 4 and 11 (17%) Stage 3 disease. Participant characteristics including HPV status, primary site and staging  and are summarised in n = 4; nodal, primary and distal metastatic recurrence n = 1; isolated primary recurrence n = 1; distal metastases alone n = 4). Five participants with nodal recurrence underwent salvage neck dissection. Median time to recurrence was 185.5  days post treatment. There were 55/65 patients with 2 year DFS and 59/65 patients with 2 year LRFS. The subsequent median follow-up was 4.1  years post-treatment and there were no other cases of progressive disease. There were 57/65 patients who attended for the 12 week post-CRT 18F-FDG PET-CT study.At 2 year follow-up, there were a total of 10 participants with recurrence , the largest lymph node (n = 8) or both locations (n = 51). Median area of aROI was 166 [88-346] mm2 for primary tumour and 281 [171-415] mm2 for the lymph nodes. Median volume of vROI was 8908  mm3 for primary tumour and 4220  mm3 for the lymph nodes. The mean cervical spinal cord ROI ADCmean was 1004 ± 79 x 10−6 mm2/s.ROI was delineated at the primary tumour . However, the the nodal ADC values did not differ between HPV OPC and other HNSCC subgroups (p = 0.3–0.61).The ADCmean and 2 year LRFS  and DFS (p = 0.02) on univariate analysis  and LRFS (p = 0.05) with odds ratio (95% CI) of 4.85 (1.18–19.8) for DFS and odds ratio (95% CI) of 5.87 (0.97–35) for LRFS ; however, due to the limited sample size 2 year LRFS analysis was not possible. Similarly, ADC values were unable to predict 2 year DFS in other HNSCC (p = 0.66–0.85), although the sample size was limited. In participants without 2 year DFS, the ADC values were markedly lower in HPV OPC compared to other HNSCC (mean at the primary tumour site was 863 ± 74 x 10−6 mm2/s for HPV OPC (n = 4) whilst 1138 ± 179 x 10−6 mm2/s for other HNSCC (n = 6), however, the sample size without 2 year DFS was too small for statistical comparison.The subgroup analysis did not demonstrate any association between HPV OPC ADC values and 2 year DFS (er HNSCC . For insp < 0.05) demonstrated between the pre-treatment ADCmean or ADCmin values and the 12 week post-CRT 18F-FDG PET-CT SUVmax when applying any of the different ROI methods at either primary tumour and nodal locations.There was no significant correlation (The ICCs for the sample of ROIs performed by two observers were 0.97 (vROI), 0.98 (aROI) and 0.98 (rROI) for the primary tumour and 0.98 (vROI), 0.98 (aROI) and 0.94 (rROI) for the lymph node locations.mean and 2 year LRFS detected on univariate analysis was no longer evident. The pre-treatment primary tumour ADC values were lower in HPV OPC than other HNSCC, both overall and in those without 2 year survival. In this study cohort, it was not possible to demonstrate any prognostic ability of pre-treatment ADC values, when stratified by HPV OPC status.This study provides data concerning the impact of HPV OPC status on the ability of pre-treatment ADC values to predict survival outcomes. On multivariate analysis, the HPV OPC status was the only independent variable predicting both 2 year DFS and LRFS, and the apparent association between the primary tumour aROI ADC6–8,11,12,15,18–21 It has been proposed that this may relate to the histological findings of higher stromal content, lower cellularity and micronecrosis, all of which are known to be associated with greater treatment resistance. However, a broad range of ADC thresholds have been reported for the prediction of treatment response and some studies have shown no significant association between the ADC values and outcome.3–5,9,10,13,16,22–25 Furthermore, one study has revealed the conflicting finding of poorer outcomes in tumours with lower ADC values; a result possibly explained by an association with poorly differentiated HNSCC.17Many previous studies have shown that an increased pre-treatment ADC is associated CRT treatment failure in HNSCC.27–30 Despite this, studies correlating ADC values with post-CRT survival outcomes generally comprise heterogenous cohorts and only a few provide data on HPV OPC status15,17 or analyse HPV OPC cohorts separately.4,13,24,35,36 There are only four studies which have considered HPV OPC status as a co-variate.4,14,15,24 Ravanelli et al24 conducted a retrospective study comparing pre-treatment ADCmean at the primary oropharyngeal tumour site in both HPV positive and negative OPC. In keeping with the results of this study, they also demonstrated that primary tumour aROI ADCmean predicted survival outcomes on univariate analysis but not on multivariate analysis.24 Cao et al4 and Martens et al14,15 adopted a similar approach to our study and compared HPV OPC with other HNSCC at both primary tumour and nodal sites. Cao et al4 concluded that pre-treatment ADC values in HPV OPC were not associated with treatment failure; however, unlike the present study, they did demonstrate a correlation in their larger cohort of other HNSCC. Martens et al14,15 performed both retrospective and prospective studies and did not find pre-treatment ADCmean to be predictive of survival outcomes in a multivariate analysis which included HPV status. The current study adds to the existing literature by analysing pre-treatment ADC values in HPV OPC and other HNSCC cohorts with a more comprehensive methodology, which included multiple approaches to contouring ROIs and the application of both ADCmean and ADCmin.Previous data also suggest that HPV OPC is associated with lower pre-treatment ADC values compared to other HNSCC, and this finding is corroborated by our data.6–8,11,12,15,18–21 has been considered counterintuitive, since low ADC values are generally found in poorly differentiated HNSCC, which would be expected to have a poorer prognosis. However, it could be postulated that the association of lower ADC values with HPV OPC status is primarily responsible for the apparent improvement in outcomes. Such a hypothesis is supported by our data, since multivariate analysis indicated that primary tumour ADCmean was no longer of prognostic significance, and it was only HPV OPC status that was an independent factor associated with 2 year survival.The favourable treatment response of low ADC tumours in previous studiesmean values in participants that did respond to treatment in HPV OPC (e.g. aROI 686–1684 x 10−6 mm2/s) would also limit the prediction on an individual basis without this knowledge.An awareness of HPV OPC status is clearly of importance in order to interpret the significance of pre-treatment ADC values. The markedly lower primary tumour ADC values in HPV OPC non-survivors compared to other HNSCC non-survivors would preclude the prediction of treatment failure without information on HPV OPC status. Similarly, the wide range of the primary tumour pre-treatment ADC24 whilst the other evaluated vROI.4,13 Our data indicate that the choice of ROI analysis may potentially influence the ability to predict survival outcomes.Another factor to consider in the interpretation of previous literature, is that different methods of contouring ROIs have been variably applied. Few studies have applied multiple methods in the same cohort and assessed the impact on the prognostic value of measurements obtained. Whilst smaller ROIs are potentially less representative, they may benefit from the ability to exclude macroscopic necrosis, so obtaining a value for the “viable tumour” and avoiding partial volume effects. Of the previous studies investigating the prognostic ability of ADC values according to HPV OPC status, one study measured aROImean and ADCmin parameters. Histological heterogeneity within tumour tissue may not be accurately reflected by the ADCmean. Whilst ADC histogram analysis is being explored,10 simple ADC parameters such as ADCmin may probe the different tumour cell populations. Since our vROI and aROI tumour contours were potentially influenced by partial volume effects from the air-filled lumen, the ADCmin was calculated as one standard deviation below the mean. However, this study did not demonstrate any benefit to using ADCmin in predicting treatment outcomes. Interestingly, Martens et al demonstrated the maximum ADC value to be a valuable prognostic pre-treatment parameter.13A further aspect of the methodology in the current study was the acquisition of both ADC2 with a wider standard deviation of ADC values.25 This introduces the potential for Type 2 errors, thus also limiting the ability of pre-treatment ADC values to predict treatment failure in the HPV OPC subgroup. However, the similarity between ADC values in HPV OPC participants with and without 2 year LRFS and DFS . The dominance of the HPV OPC subgroup led to a smaller number of HNSCC participants which limited the scope of robust statistical comparisons with survival outcomes. Moreover, it is recognised that HPV OPC is associated with a lower rate of treatment failure than the 30% applied to the power calculation and DFS  would armean and 2 year LRFS, was not confirmed on multivariate analysis of all participants or in the subgroup analysis of the HPV OPC participants. A potential explanation for this is that the HPV OPC status, which is widely accepted to carry a favourable prognosis, also happens to demonstrate lower ADCmean values. Larger cohorts with separate analyses of subgroups stratified by HPV-OPC status are required in order to advance our understanding of the optimal utilisation of DW-MRI in the evaluation of treatment responses in HNSCC.In conclusion, this comprehensive methodology adds to the recent data concerning the impact of HPV OPC status on the ability of pre-treatment ADC values to predict survival outcomes. The apparent association between lower pre-treatment primary tumour ADC


Pseudoachondroplasia (PSACH), a short limb skeletal dysplasia associated with premature joint degeneration, is caused by misfolding mutations in cartilage oligomeric matrix protein (COMP). Here, we define mutant-COMP-induced stress mechanisms that occur in articular chondrocytes of MT-COMP mice, a murine model of PSACH. The accumulation of mutant-COMP in the ER occurred early in MT-COMP articular chondrocytes and stimulated inflammation (TNFα) at 4 weeks, and articular chondrocyte death increased at 8 weeks while ER stress through CHOP was elevated by 12 weeks. Importantly, blockage of autophagy (pS6), the major mechanism that clears the ER, sustained cellular stress in MT-COMP articular chondrocytes. Degeneration of MT-COMP articular cartilage was similar to that observed in PSACH and was associated with increased MMPs, a family of degradative enzymes. Moreover, chronic cellular stresses stimulated senescence. Senescence-associated secretory phenotype (SASP) may play a role in generating and propagating a pro-degradative environment in the MT-COMP murine joint. The loss of CHOP or resveratrol treatment from birth preserved joint health in MT-COMP mice. Taken together, these results indicate that ER stress/CHOP signaling and autophagy blockage are central to mutant-COMP joint degeneration, and MT-COMP mice joint health can be preserved by decreasing articular chondrocyte stress. Future joint sparing therapeutics for PSACH may include resveratrol. Cartilage oligomeric matrix protein (COMP) is a large, matricellular protein that mediates a variety of cell–cell and cell–matrix interactions ,4,5,6,7.Long before COMP mutations were identified as the cause of PSACH, it was known that growth plate chondrocytes contained massive amounts of material in the rER cisternae. Later studies proved that mutant-COMP does not fold properly and, therefore, is retained in the ER ,39. MoreIn this work, we focused on delineating the effect of mutant-COMP on joint health in our PSACH mouse model and the timing of joint degeneration. Articular and growth plate cartilages serve very different functions, and these tissues mature at different times ,47. GrowIntracellular retention of mutant-COMP in chondrocytes is a hallmark of PSACH ,40,41. PIn growth plate chondrocytes, the intracellular accumulation of mutant-COMP stimulates a complex pathological process. This involves activation of the ER stress through the CHOP pathway, which in turn activates oxidative and inflammatory processes that exacerbates ER stress, causing an over activation of mTORC1 signaling blocking autophagy, and ultimately chondrocyte death ,44. As s8 ± 6.52 × 107) than in the controls (3.28 × 108 ± 1.88 × 107) (p < 0.034911) (data not shown). These findings show that joint degeneration-associated cytokines (TNFα beginning at 4 weeks) and elevated MMP13 immunostaining were associated with mutant-COMP accumulation in articular chondrocytes in MT-COMP mice.Because inflammatory processes involving pro-inflammatory cytokines, IL-1β and TNFα, play a significant role in the growth plate, these cytokines were investigated in the articular cartilage (IL-1β data not shown). Shown in Autophagy is repressed in both OA articular chondrocytes  and MT-CSenescence is known to play a role in OA joint degeneration ,55 and wIn mice, the presence of pain is measured by changes in specific behaviors . One sucAlterations of gait can indicate the presence of pain because these alteration are a natural attempt to reduce stress on limbs associated with pain during ambulation . The DigOA scoring was used to detect and quantify early degenerative changes in the proteoglycan content of articular cartilage, synovitis, and bone/cartilage. No differences were detected prior to 16 weeks of age. MT-COMP femur proteoglycan content was less, and there was a trend toward higher scores for bone/cartilage damage and total score at 16 weeks. . A totalp < 0.0005) similar to controls 2.56 ± 2.15 (data not shown). Importantly, resveratrol treatment prevented the accumulation of mutant-COMP in the ER of articular chondrocytes , inflammation (TNFα), ECM degradative enzymes (MMP-13), block of autophagy (pS6), and senescence (p16 INK4a). Importantly, these stresses were not observed in MT-COMP mice in the absence of DOX . TNFα stAging and/or cellular stress stimulates senescence ,66, and The timing of mutant-COMP joint degeneration is distinct from the mutant-COMP growth plate pathology. In articular chondrocytes, mutant-COMP retention and inflammation start at 4 weeks, and ER stress, blockage of autophagy, and chondrocyte death are seen between 8 and 12 weeks (summarized in MT-COMP mice undergo premature joint degeneration far earlier than the background strain (C57BL\6) that develop joint degeneration beginning at 1 year of age. While there is limited information about the temporal development of joint degeneration in PSACH, natural history studies show that there is premature joint degeneration in PSACH, starting in mid- to late teenage years . Based o−/−mice. Notably, the MT-COMP/CHOP−/−mice still express mutant-COMP, but in the absence of CHOP the mutant-COMP pathology is diminished. Similarly, resveratrol treatment from birth to 20 weeks dampens the mutant-COMP pathologies of ER stress (CHOP), inflammation (TNFα), block of autophagy (pS6), and senescence (p16 INK4a). This reduction of ER stress and inflammation interrupts the pathological loop between ER, oxidative stress, and inflammation. Moreover, resveratrol restoration of autophagy allows clearance of mutant-COMP from the ER of articular chondrocytes. These proof-of-principle findings show that the articular cartilage of MT-COMP mice can be preserved if intervention occurs prior to a point of no return and opens new therapeutic avenues for PSACH.The essential role of CHOP in the mutant-COMP joint degenerative process is illustrated by the alleviation of articular chondrocyte stress in MT-COMP/CHOPThe MT-COMP mice used in these and previously described experiments contain the pTRE-COMP (coding sequence of human COMP + FLAG tag driven by the tetracycline responsive element promoter) and pTET-On-Col II (rtTA coding sequence driven by a type II collagen promoter) ,47,53. D−/−), as used in previous experiments [CHOP null mice were procured from Jackson Laboratories and mated with MT-COMP bigenic mice to obtain a strain expressing MT-COMP in a CHOP null background . Based on this information, 5 mice/group were analyzed because OA gait dysfunction is subtler to detect compared to ethanol induced ataxia. ANOVA analysis was performed, followed by pairwise comparisons with Kruskal–Wallis and post hoc Dunn’s analysis. All mice were moved to the behavior room at 8:00 a.m. and DigiGait analysis occurred between 9:00 a.m.–12:30 p.m. Due to the dwarfing phenotype associated with expression of mutant-COMP in mice [Gait of male mice was analyzed using a DigiGait treadmill system  following the protocol described for collagen-induced OA . Previou in mice ,46,47, o in mice  cannot bOA scoring was performed on 10 different 5 um sagittal sections . To ensure sections were from the same area of the joint; only sections that contained both menisci were scored. While OARSI scoring covers a wide range of OA pathology, in this study, OA scoring was modified to optimize evaluation of early OA pathology. Four areas, synovium, bone/cartilage, tibial and femoral articular cartilage, were scored from 0 to 3 on each safranin-O stained section. A score of 0 indicated normal or no damage, 1 = mild damage, 2 = moderate damage, and 3 = denotes severe damage. Synovitis, bone/cartilage damage, and proteoglycan of the tibia and femur were scored individually, and all scores were summed with maximal damage being associated with a score of 12. Synovitis was defined as: mild—increase in thickness of synovial lining and increase in stromal area, moderate—increase in stromal density, or severe—thickening of synovial lining with further increase of stromal cellular density. Bone/cartilage damage was defined as: normal—surface was smooth, mild—minor erosion of the surface, moderate—presence of remodeling with minor erosion, or severe—major erosion. Proteoglycans of the articular cartilage of the tibia and femur was classified as: normal—if staining was even through to the subchondral bone, mild—when staining was thinned, moderate—thinning of proteoglycan stained layer and absence of staining in some areas, or severe—widespread loss of proteoglycan staining. Ten mice per experimental group were used for each time point, providing 80–90% power to detect a minimal difference of 2 or 3 units. All scoring was performed blindly. Section depth, thickness, fixation, and decalcification conditions were all identical for all limbs analyzed. The Kruskal–Wallis test was used to evaluate distribution of OA scores across 6 experiment groups, with Post-hoc Dunn’s test comparing MT-COMP to controls.https://www.perkinelmer.com/lab-solutions/resources/docs/APP_Protocol_MMPSense%20680.pdf, accessed on 22 May 2021). MMP 680 is an optically silent substrate that fluoresces when cleaved by MMP-2, -3, -9, and -13. MMPSense signal was assessed blindly by positioning a circle of a standard size around the knee , and radiance efficiency was generated from IVIS software. Six mice were included per group, and the system had the power to detect a difference of 30% or greater. Mann–Whitney U test was used to evaluate MMP activity in control and MT-COMP mice.Mice were treated with depilatory cream to remove hair prior to imaging. MMP 680 reagent was injected into the tail vein, and animals were imaged 24 h later, as per manufacturer’s instructions, along with an uninjected control (UIC). Males were imaged on an IVIS Spectrum In Vivo Imaging System  (


The process evaluation of the program was positive. The CBO-private clinic service model was helpful in increasing HPV vaccination uptake among MSM.This study evaluated the effectiveness of the community-based organization (CBO)-private clinic service model in increasing human papillomavirus (HPV) vaccination uptake among unvaccinated men who have sex with men (MSM) in Hong Kong during a 12-month follow-up period. A CBO-private clinic model was implemented to promote HPV vaccination among Chinese MSM. A CBO with good access to MSM approached MSM aged 18–45 years who had never received an HPV vaccination, invited them to receive an online health promotion, and referred them to receive HPV vaccination at gay-friendly private clinics. A baseline survey and a follow-up evaluation at Month 12 were conducted. A total of 350 participants completed the baseline survey. Among 274 participants who were followed up at Month 12, 46 (16.8%) had taken up at least one dose of HPV vaccination. After adjusting for significant baseline characteristics, the perceived susceptibility (AOR:1.25,  Across countries, men who have sex with men (MSM) have a much higher risk of contracting human papillomavirus (HPV) and its related diseases  than the general male population ,2,3,4. AHPV vaccination is highly effective in preventing vaccine-type genital warts and cancers among MSM ,16 and oThe CDC recommends MSM aged ≤45 years to receive the HPV vaccination ,22. In tA randomized controlled trial (RCT) was conducted between 2017 and 2019 among MSM in Hong Kong to evaluate online intervention promoting HPV vaccination . The resCommunity-based organization (CBO) in Hong Kong has a good position of promoting HPV vaccination among local MSM. First, half of MSM usually received HIV prevention services provided by local CBO  ,33. SecoSuggestion from peer CBO workers can serve as a strong cue to action . MoreoveThis longitudinal study was conducted from August 2019 to April 2021. All participants completed a telephone survey before they received online interventions promoting HPV vaccination, and completed another telephone survey 12 months after the baseline survey. There was no control or comparison group. The flowchart diagram was shown in Inclusion criteria were: (1) Hong Kong Chinese speaking males aged 18–45 years old, (2) self-reported having had oral or anal intercourse with at least one man in the last six months, (3) never received HPV vaccination, and (4) willing to complete a follow-up telephone survey 12 months after the baseline survey.Trained staff of a CBO providing HIV-related services recruited participants through multiple sources, including outreaching in gay bars or saunas in Hong Kong, online recruitment, and referrals made by peers and other CBOs. Detailed recruitment procedures were reported in a published paper . InteresBy appointment, the CBO staff conducted the 15-min baseline survey by telephone. All participants received the following health promotion components:(1) Watching an online video promoting HPV vaccination for MSM: The online video developed by our team in a previous RCT was modified for this study . The modIn the video, a peer MSM talked about the risk of having penile or anal cancers among MSM, the severe consequences of HPV-related diseases, high efficacy, and long protection duration of HPV vaccination. The video also contained flashes of scary images of genital warts and penile and anal cancers to increase the perceived severity. The peer MSM also emphasized that HPV vaccination was a worthy long-term investment for their health. He demonstrated the procedures for receiving HPV vaccination in one of the collaborative private clinics, which was convenient, caring, privacy guaranteed, and non-judgmental. Participants could not fast-forward or skip any part of the video.(2) Each participant was sent a discount coupon through SMS/WhatsApp and could enjoy a 10% discount for taking up three doses of HPV vaccines at a collaborative private clinic.(3) Participants could access the project webpage by scanning the Quick Response (QR) code on the discount coupon. Through the webpage, participants could: (i) watch the aforementioned online video; (ii) read the description of the project, knowledge, and benefits of HPV vaccination, information about HPV/HPV-related diseases that increased the perceived susceptibility and perceived severity; (iii) access a discussion forum containing positive feedbacks of peers who have taken up HPV vaccination. These testimonials provided cue to action supporting HPV vaccination; and (iv) obtain the contact information of the project staff and information about the collaborative private clinics.(4) Five reminders were sent to participants by SMS/WhatsApp at Month 1, 2, 4, 6, and 8.During the project period, two types of HPV vaccines were available for males . For participants who decided to take the vaccine in the collaborative private clinics, they could sign up on the project webpage by filling up their contacts and preferable timeslots. The CBO staff would contact them by telephone or WhatsApp to facilitate appointment making to take up HPV vaccination at collaborative private clinics at a 10% discount from the market rate (about HK$4200 or US$542 for three doses). Participants could also directly contact the collaborative clinics to make an appointment. The project did not limit participants’ choice for taking up HPV vaccination in other places.Participants were invited to complete a follow-up telephone survey 12 months after the baseline survey. Up to five calls were made at different timeslots during weekdays and/or weekends before considering a participant as a dropout.The primary outcome of the study was the validated uptake of any doses of HPV vaccination within a 12-month follow-up period. To validate HPV vaccination uptake, the participants were requested to send the project staff an image of their receipt without personal identification after they received each dose of HPV vaccines. The same approach to validate HPV vaccination uptake was used in a previous study . VaccinaVarious baseline information was collected, such as socio-demographics, sexual orientation, HIV or STI prevention service utilization, history of HIV and other STIs, and queried sexual behaviors, which included anal intercourse with male regular and non-regular sex partners and male sex workers, condomless anal intercourse (CAI) with men, multiple male sex partnerships, sexualized drug use (SDU), and so on. Regular male sex partners (RP) were defined as lovers and/or stable boyfriends, while non-regular male sex partners (NRP) were defined as casual sex partners. In this study, SDU was defined as the use of any psychoactive substance before or during sexual intercourse ,35.Six items measured knowledge related to HPV and HPV vaccination. A composite variable was constructed by counting the number of correct responses (ranged from 0 to 6). Five scales assessed perceptions based on the HBM, including a three-item Perceived Susceptibility Scale, four-item Perceived Severity Scale, four-item Perceived Benefit Scale, five-item Perceived Barrier Scale, two-item Cue to Action Scale, and two-item Perceived Self-Efficacy Scale. These scales were modified from those used in a previous study among MSM in Hong Kong .At Month 12, participants were asked to evaluate the health promotion . Participants who have taken up HPV vaccination were asked additional questions rating their satisfaction of the services provided by the collaborating clinics.The difference in baseline characteristics between participants who completed Month 12 follow-up and dropouts were compared by using chi-square tests  or Mann–Whitney U tests (for continuous variables). The subsequent analysis was performed with data from those who had completed both surveys. We used the validated uptake of any doses of HPV vaccination during the follow-up period as the dependent variable and baseline background characteristics  as independent variables. p < 0.05 in the univariate analysis, the association between knowledge and perceptions related to HPV and HPV vaccination and the dependent variable were then assessed by adjusted odds ratios (AOR). Each AOR was obtained by fitting a single logistic regression model, which involved one of the perceptions and the significant background variables. SPSS version 21.0  was used for data analysis, and p < 0.05 (two-sided) is considered as statistically significant.Crude odds ratios (OR) predicting that the dependent variable were obtained using logistic regression models. After adjustment for those variables with Four hundred of the 565 prospective participants were screened to be eligible, 50 of whom refused to participate in the study for time or other logistic reasons, and 350 (87.5%) completed the baseline survey and received health promotion. At Month 12, 274 (78.3%) completed the follow-up survey.At baseline, about half of the participants were 18–30 years old (50.3%), and with a monthly personal income at least HK$20,000 (US$2565) (57.4%). The majority of the participants were currently single (75.7%), had attained tertiary education or above (86.6%), were employed full-time (82.9%), and identified themselves as homosexuals (91.1%). Among the participants, 1.7%, 8.9%, and 16.9% reported a history of HIV, HPV infection/genital warts, and other STI infection, respectively. In the past six months, 50.3% and 54.6% reported CAI with men and multiple male sex partnerships. p = 0.02), anal intercourse with NRP (p = 0.02), and male sex workers (p = 0.01), no significant difference was found between participants who completed the Month 12 follow-up and dropouts . Single factors for these scales by exploratory factor analysis, which explained 56.2–82.5% of the total variance. Apart from self-reported HIV sero-status . All of them were able to provide receipts for verification. The location for receiving HPV vaccination included the collaborative private clinics  and other private hospitals/clinics . The participants self-paid HK$2000–6800  to receive the vaccination. Most vaccinated participants reported no side effects . The reported side effects included pain at the injection site , fatigue , and dizziness . Most of these side effects were very mild or mild .At Month 12, 46 participants self-reported having had taken up at least one dose of HPV vaccination (three doses: p < 0.001) and utilization of HIV/STI prevention services other than HIV testing at baseline  were associated with higher HPV vaccination uptake during the follow-up period , (2) perceived consequences of HPV-related diseases to be severer , (3) perceived benefits of HPV vaccination , and (4) perceived self-efficacy of taking up HPV vaccination . In addition, behavioral intention to take up HPV vaccination at baseline was also associated with higher uptake during the follow-up period   perceived higher risk of contracting HPV and HPV-related diseases (perceived susceptibility)  .At Month 12, 228 unvaccinated participants were asked about their reasons for not taking up HPV vaccination. The most commonly mentioned reason was the high cost of HPV vaccination , followed by feeling unnecessary to receive such vaccination , concerns about COVID-19 transmission , and no time to do so .Among 274 participants who completed the Month 12 survey, 72.3% and 38.7% believed the health promotion video was clear and attractive respectively. The participants thought the video was helpful in the following aspects, such as increasing awareness of benefits of HPV vaccination (73.4%), reducing barriers to receive HPV vaccination (46.0%), and increasing self-efficacy (56.2%) and intention (40.5%) to receive HPV vaccination.Among 46 vaccinated participants, 50–95.7% rated “satisfactory” to the following services/features of the collaborating clinics: (1) convenience of the location of clinic (89.1%), (2) convenience of the arranged time slot for vaccination (95.7%), (3) waiting time between making appointment and receiving the first dose (84.8%), (4) level of privacy (69.6%), (5) staff’s acceptance of MSM’s subculture (50.0%), (6) explanation made by the staff of private clinics (73.9%), and (7) professionalism of the staff of private clinics (76.1%) .It is important to transform research findings into regular services. We involved CBO and private clinics as key stakeholders at the planning stage. The implementation was smooth and concurred with routine services of CBO and private clinics. The evaluation results showed that the CBO-private clinic service model was helpful in increasing HPV vaccination uptake among MSM in Hong Kong, as 16.8% of participants being followed up at Month 12 had taken up at least one dose of HPV vaccination. We expected most of these participants would complete all three required doses in near future, as all of them settled the payment for the entire package when receiving the first dose and made appointments for receiving the remaining doses. Although there was an increase in the cost of HPV vaccines (from HK$3800 or US$490 to HK$4200 or US$542) and there was no MI, the uptake rate was comparable to the intervention group (17.3%) in previous RCT . HoweverThe findings also had some implications to health service and health care policy. Our results supported that HPV vaccination was accepted by local MSM at the market rate. Providing subsidized or free HPV vaccination would further increase the uptake rate. MSM in Australia and the United Kingdom responded well to pilot programs providing free HPV vaccination ,24,25. HThe planning of future programs can be facilitated by the experiences of the present study. We found that those with history of HPV infection and/or genital warts were more responsive to the health promotion as compared to MSM without such history. It is possible that MSM with such history would perceive a stronger need to protect themselves by taking up HPV vaccination. Future studies are needed to explore whether different strategies should be applied for MSM with and without prior experience of HPV infection/genital warts. We also found that the baseline measurement of perceived susceptibility, perceived severity, perceived benefit, perceived self-efficacy, and behavioral intention related to HPV vaccination significantly predicted HPV vaccination uptake during the project period. Since our health promotion was standard and one-off, such findings suggested that some follow-ups should be implemented to modify these perceptions. Meta-analysis suggested that interventions tailored to one’s stage of change (SOC) were more effective than non-stage-tailored, especially among less motivated individuals . StudiesMoreover, asking people to provide where, when, and how they want to perform a behavior could increase the perceived self-efficacy . The resThis study had the strengths of a relatively low dropout rate, validated primary outcome, and good process evaluation. However, it had several limitations. First, there was no control or comparison group. The aim of this study was to evaluate the effectiveness of the service model in real-world setting. Second, a selection bias existed, as we were not able to collect information from MSM who refused to join the project. They might have different characteristics comparing to participants. Third, attrition bias existed. The dropouts were more likely to be HIV positive or with unknown sero-status and had anal intercourse with NRP and male sex workers at the baseline. However, the bias should be limited as these baseline characteristics did not significantly predict HPV vaccination uptake during the project period. Fourth, we did not ask for details about how COVID-19 influenced the participants’ HPV vaccination. However, a previous study exploring the difficulties to access HIV-related services among MSM in Hong Kong during the same period provided some insights. Worrying about being exposed to a potentially COVID-19 infectious environment, experiencing disruptions in work due to COVID-19 and its control measures, and reduced connection to the MSM community during the pandemic were associated with increased difficulties in accessing HIV-related services in general . Some ofThe CBO-private clinic service model was helpful in increasing HPV vaccination uptake among MSM in Hong Kong. A larger-scale pilot program should be considered based on the experience of this project.


Triticum aestivum L.) is one of the most important cereal food crops for the global population. Spike-layer uniformity -related traits (SLURTs) are quantitative and have been shown to directly affect yield potential by modifying the plant architecture. Therefore, these parameters are important breeding targets for wheat improvement. The present study is the first genome-wide association study (GWAS) targeting SLURTs in wheat. In this study, a set of 225 diverse spring wheat accessions were used for multi-locus GWAS to evaluate SLURTs, including the number of spikes per plant (NSPP), spike length (SL), number of spikelets per spike (NSPS), grain weight per spike (GWPS), lowest tiller height (LTH), spike-layer thickness (SLT), spike-layer number (SLN) and spike-layer uniformity (SLU).Bread wheat  were identified when the analysis was both performed individually and combined for two environments. Twenty-nine MTAs were detected in environment one, 48 MTAs were discovered in environment two and 59 MTAs were detected using combined data from the two environments. Altogether, 15 significant MTAs were found for five traits in one of the two environments, and four significant MTAs were detected for the two traits, LTH and SLU, in both environments i.e. E1, E2 and also in combined data from the two environments. In total, 279 candidate genes (CGs) were identified, including Chaperone DnaJ, ABC transporter-like, AP2/ERF, SWEET sugar transporter, as well as genes that have previously been associated with wheat spike development, seed development and grain yield.The MTAs detected through multi-locus GWAS will be useful for improving SLURTs and thus yield in wheat production through marker-assisted and genomic selection.The online version contains supplementary material available at 10.1186/s12864-021-07834-5. Triticum aestivum L.) is one of the most widely grown cereal crops in the world and serves as the main energy source for approximately one-third of the global population . The results were compared based on the physical position of the particular marker on a particular chromosome for the same trait or grain yield. Representative chromosomal maps were prepared for the comparative presentation of all MTAs detected in the present study and previously reported QTLs/MTAs for each trait on an individual chromosome using Map chart software [The MTAs identified were also compared with previously reported QTLs/MTAs for the same traits and grain yield. The physical positions of all MTAs detected in the present study and previously reported QTLs/MTAs were identified through Ensembl Plants [version 50; software .https://www.rproject.org/contributors.html).The GS method cBLUP implemented in GAPIT version 3 was used to obtain GS values across the measured traits. Graphics of GS were generated in the R statistical environment using the base package and functions . High-confidence annotated genes were retrieved from a chromosome-wise LD decay distance window for each identified MTA. This window size varies for each MTA depending on the local LD decay for each chromosome. The gene ontology (GO) annotation information of these candidate genes (CGs) was extracted from the IWGSC website (http://www.wheatgenome.org/). In silico gene expression analysis was also conducted to identify CGs using RNA Seq expression data from Wheat Expression Browser (http://www.wheat-expression.com/). A heatmap was generated for the presentation of expression data of genes in different stages and tissues considering the selected domain relevant to the studied traits.A total of 136 significant MTAs were identified in E1, E2 and the combined data, out of which 94 unique MTAs were subjected to the identification of CGs for all the study traits  related traits (SLURTs) shown with wheat plants having similar tillers height (a) and different tiller height (b). In (a) only one spike layer can be seen from the vertical perspective (SLN = 1) as spikes per plant have consistent vertical distribution (SLU = 1) and the SLT was identical to one spike length (SL) whereas in (b) two spike layer can be seen from the vertical perspective (SLN = 2) as spikes per plant have inconsistent vertical distribution (SLU = 0.5) and the SLT was identical to two spike length. A detailed description of SLURTs is provided in Table Fig. S2. Pairwise Pearson correlation among the eight SLURTs in E1 and grain yield (GYPP). A single asterisk (*) represents 0.05 level of significance. A double asterisk (**) represents 0.01 level of significance and a triple asterisk (***) represents 0.001 level of significance. Fig. S3. Pairwise Pearson correlation among the eight SLURTs in E2 and grain yield (GYPP). A single asterisk (*) represents 0.05 level of significance. A double asterisk (**) represents 0.01 level of significance and a triple asterisk (***) represents 0.001 level of significance. Fig. S4. Pairwise Pearson correlation among the eight SLURTs and grain yield (GYPP) for the combined E1 and E2 environments. A single asterisk (*) represents 0.05 level of significance. A double asterisk (**) represents 0.01 level of significance and a triple asterisk (***) represents 0.001 level of significance. Fig. S5. Manhattan plots obtained after using FarmCPU for eight SLURTs (a) GWPS (b) LTH (c) NSPP (d) NSPS (e) SL (f) SLN (g) SLT (h) SLU under E1 (above numbers) and E2 (below numbers) conditions. Numbers correspond to wheat chromosome. Fig. S6. QQ-plots to visualize the deviation of observed p values from expected p values (based on null hypothesis) for eight SLURTs (a) GWPS (b) LTH (c) NSPP (d) NSPS (e) SL (f) SLN (g) SLT (h) SLU under E1 and E2 conditions. Fig. S7 (i-v). Comparison of MTAs or MTA groups, detected in the present study, with historical QTLs/MTAs in different chromosomes; MTAs detected in known flanking regions or locations of QTLs/MTAs for same traits or grain yield are depicted in green color and MTAs found near to flanking regions or locations (less than 50 Mb) of QTLs/MTAs depicted in red colour. Flanking markers are shown with the same colour and with the reference details of the previous study. Novel MTAs detected in the present study are presented without any colour. The corresponding physical distances (Mb) of the QTL/MTA regions on each chromosome were obtained by blasting the flanking sequences of markers, depicted on the right side of each figure, to the Chinese Spring RefSeq v1.0. GY; grain yield, TN; tiller number.Additional file 2: Table S1. Descriptive statistics details for all the studied traits. Table S2. Phenotypic data of environment 1 (E1: 2017–18), environment 2 (E2: 2018–19) and mean of E1 and E2. Table S3. List of candidate genes identified using MTAs detected in E1, E2 and combined (E1 and E2).