Aim: to evaluate SNHL related to CSOM and its association with cholesteatomas, disease duration and patients’ ages. Methods: Retrospective analysis of 115 patients with CSOM with and without cholesteatoma submitted to surgical treatment. Inclusion criteria were active unilateral disease, normal contralateral ear and age below 60 years. Results: The average age was 26.3 years, 58 males and 57 females. The duration of ear disease was, in average, 12.4 years. The average threshold of hearing was 40 dB in CSOM ear and 22 dB in the normal contralateral ear (P=0.002). CSOM with cholesteatoma occurred in 78 of 115 cases. In the abnormal ear, SNHL was seen in 15 cases, being 6 cases of profound loss, that correlated with adjusted-age (P=0.003) and absence of cholesteatoma (P=0.01), but not with disease duration (P=0.458). Conclusion: SNHL occurred in 13% of the patients with CSOM, and was correlated with older age, but not with the presence of cholesteatoma or longer duration of ear disease.Sensorineural hearing loss (SNHL) related to chronic suppurative otitis media (CSOM) was studied to clarify the involvement of cholesteatomas in this context.  Chronic otitis media may be considered a public health problem. In the United States, it is estimated that over two billion dollars are spent yearly to treat both acute and chronic ear infectionsNow, considering suppurative chronic otitis media (SCOM), this may or may not be associated with the development of a cholesteatoma, and conductive hearing loss is the characteristic audiologic alterationThe association between chronic otitis media and sensorineural hearing loss (SNHL) has been broadly studied and remains a controversial topic. Studies present a frequency that varies from no clinical significanceThe goal of the present investigation was to correlate SCOM associated with SNHL with cholesteatoma with patient’s age and disease duration.The present study was carried out through the analysis of charts from those patients submitted to otologic surgery because of SCOM with or without cholesteatomas. As for inclusion criterion we considered chronic unilateral otorrhea with a normal contralateral ear in patients with less than 60 years of age. As exclusion criteria, we considered the family history of congenital or acquired SNHL, history of noise exposure, head injury, prior ear surgery, or the possibility of a perilymphatic fistula. The socio-economic statuses of the patients were controlled: having a private health insurance and having been submitted to ear surgery in a hospital that does not belong to the public health care system.All patients were assessed, treated and followed up by the same professional, with vast experience in otology. In the month prior to the otologic surgery, audiometry exams were carried out in double by two independent professionals and who did not know about the patient’s clinical history or about the presence or absence of cholesteatoma. The exams were carried out in the same equipment and in a sound treated booth. In the audiometry analysis, the air-bone gap considered was of 10 dB to define a conductive hearing lossWe assessed the correlation between SNHL with the following variables: age, ear disease duration and the presence or absence of cholesteatoma. The criterion to define the presence of cholesteatoma was the pathology description of the surgical specimen. In the case of the non-cholesteatomatous disease we considered the criteria of continuous otorrhea, resistant to antibiotic treatment for at least two months, associated to central eardrum perforation, and confirmation of cholesteatoma-free situation during surgery.The size of the calculated sample was of at least 95 patients, considering the SNHL frequency in patients with SCOM as being of 10%The charts of 115 patients were assessed. Mean age was of 26.3 (SD=15.66), varying between 4 and 59 years, being 58 men and 57 women.Average hearing threshold was of 40 dB in the ear with SCOM and 22 dB in the normal contralateral ear .Mean duration of the ear disease was of 12.4 (SD=10.9), varying from 1 to 42 years. In the ear with SCOM, we observed normal air conduction audiometry in 25 (21.7%) cases, mild hearing loss in 43 (37.4%), moderate in 41 (31.7%) and severe or profound in 6 (5.2%) cases. Considering the bone via, SNHL was seen in 15 (13%) cases, 8 of isolate loss in 4 KHz and 7 with losses in 0.5, 1, 2, and also in 4 KHz.In the contralateral ear with normal otoscopy (paired control), the air pathway was normal in 100 (86.9%) cases, mild in 13 (11.3%) and moderate in 2 (1.3%) cases.Considering bone conduction, we observed 5 (4.3%) cases of mild SNHL isolated in 4 KHz.Analyzing bone conduction hearing loss alone in 4 KHz, there was no statistically significant difference between the ears with SCOM and the normal contralateral ear . When we compared the alteration in the hearing threshold by bone conduction for the mean frequencies of 0.5, 1, 2 KHz in both groups, we did not see losses in the normal ear and in the ear with SCOM, moderate SNHL was seen in one case and severe to profound hearing loss in 6 cases.As to the presence of cholesteatoma, it was seen in 37 (32%) and it did not happen to 78 (68%) cases. As to the cholesteatoma interference analysis in SNHL development, In the present investigation, average hearing loss of 40 Db was higher than the one observed in other series of cases, of which average varied from 19 to 33 Db in the frequencies of speechIn the ear with SCOM, the frequency of SNHL was of 13% (15 cases). In a similar study, carried out with patients with a similar socio-economical status to those from the patients selected in the present study, SNHL was seen in 10% of the cases reviewedDisease duration does not correlate with SNHL . In a stSNHL was seen in SCOM with and without cholesteatomas . In manyAccording to the present study, SCOM with or without cholesteatoma may evolve towards SNHL, which was observed in 13% of the cases and is correlated with an age increase, but not with the presence of cholesteatoma or with a longer duration of otologic disease.


Noise-induced hearing loss (NIHL) is the most prevalent occupational disease in the world and firefighters are at increased risk of NIHL due to their frequent exposure to hazardous levels of noise during service. Adverse effects of NIHL include acceleration of age-related hearing loss and an increased risk of cognitive decline. A critical challenge in addressing NIHL is the delayed clinical presentation of symptoms and lack of sensitive tools for early detection. To study the early clinical symptoms of NIHL in this high-risk group, we collected hearing function data including behavioral audiometric thresholds and distortion product otoacoustic emissions (DPOAEs) in 176 firefighters during annual physical assessments. Results revealed significant deficits in cochlear outer hair cell function in the presence of normal audiograms. Additionally, 55% of firefighters self-reported changes in hearing, while 20% self-reported concerns about their balance. This study is the first to characterize DPOAEs in firefighters who display decreased DPOAE amplitudes with increasing years in the fire service. These effects were observed even when controlling for hearing loss and age and are suggestive of a link between hearing loss and occupational exposure to hazardous noise. Noise exposure is the leading cause of acquired hearing loss, a significant global health burden affecting nearly 500 million people worldwide . OverexpNoise-induced hearing loss (NIHL) is the most prevalent occupational disease in the world, with an estimated 1.3 billion people suffering from hearing loss due to noise overexposure . Noise eOtoacoustic emissions can be used to monitor early changes in the micromechanical function of the cochlear OHCs arising from noise exposure ,20,21,22Noise overexposure can also damage the vestibular periphery . PrecliOccupational groups such as firefighters are at particularly high risk of noise-induced changes to the inner ear due to chronic exposure to hazardous noise levels ,37,38,39Inner ear structural changes can occur years before becoming clinically evident ,3,4,5, cThis study was designed as a prospective population-based cohort study. All study procedures were approved by the University of Miami Institutional Review Board (IRB# 20200222).We collected self-reported data from 220 South Florida firefighters from a custom 53-item questionnaire on perceptions of noise exposure, hearing and balance health, and noise safety practices. As part of the survey, the demographic data collected were respondent age, gender, race, ethnicity, marital status, and education. All participants were also asked about hearing and balance health history, cognitive symptoms, exposure to noise, and use of hearing protection. Additional information reported by the participants included their current role as a firefighter, military service, occupational and recreational noise exposure, balance and auditory symptoms following exposure to hazardous noise, and use of a hearing assistive device. We further collected functional hearing outcomes in a subgroup of 176 career firefighters recruited from South Florida fire departments during their 2020 annual health examinations and present the data from this cohort here.Auditory measurements were conducted onsite at a local fire station in Miami-Dade County, Florida, during annual physical assessments of fire service members. All measurements were performed by trained research personnel in an enclosed room, free of interference from external noise. Noise levels were measured intermittently throughout testing to ensure that ambient noise did not exceed 45 A-weighted decibels (dBA), thereby meeting the ANSI S3.1–1999 standard for maximum permissible ambient noise levels for audiometric and DPOAE testing ,45.Otoscopy was performed in all participants prior to audiologic assessment. Visual examination of the ear canal and tympanic membrane was conducted to ensure normal anatomy and that external canals were free from occluding debris.Behavioral hearing thresholds were measured using air-conducted pure-tones under noise-reduction insert headphones  for both ears using an MA40 portable diagnostic audiometer  calibrated to meet the American National Standards Institute standards. Pure-tone air conduction hearing thresholds were obtained for test frequencies of 0.5, 1, 2, 3, 4, 6, and 8 kHz in 5-decibel (dB) steps using a modified Hughson–Westlake procedure . Normal 2/f1) of 1.22, L1 = 65 dB sound pressure level (SPL), L2 = 55 dB SPL, with f2 frequencies ranging from 1.5 to 10 kHz. An in-the-ear calibration provided by the Bio-Logic test equipment was utilized prior to each measurement. The probe was coupled to the participant’s ear with a disposable foam ear tip. The DPOAE response was considered present when the DPOAE amplitude was greater than 6 dB above the noise floor [DPOAEs were recorded bilaterally to objectively measure cochlear OHC function. DPOAE amplitude was measured using a Bio-Logic Scout AuDX system . Responses were elicited with two simultaneously presented primary frequency tones with a fixed primary ratio  Do you have concerns about your hearing ? (2) Do you have ringing or buzzing in the ear? (3) Do you experience changes in hearing following loud noise exposure? (4) Do you experience ringing/buzzing in your ear following loud noise exposure? (5) Do you experience muffled hearing following loud noise exposure? Responses for self-reported symptoms were collected on a five-point scale . For descriptive analysis of self-reported changes in auditory function and DPOAE findings, responses were converted to yes/no binary variables where often–always was considered ‘yes’ and never–sometimes was considered ‘no’.p-values < 0.05. Pairwise comparisons with Bonferroni corrections for multiple comparisons were used to follow up significant main effects. All analyses were performed using the SPSS® software .Descriptive analyses were based on non-missing responses and conducted for demographic variables and the 5 self-reported outcomes. Correlation analyses were used to analyze associations between self-reported changes in hearing function and DPOAE amplitude. Analyses of variance were conducted to examine the relationships between population factors and hearing function. Statistical significance was defined by Participants ranged in age from 21 to 59 years (mean 38.8 ± 8 years), years of service ranged from 1 to 31 years (mean 12.3 + 7 years), and 95% identified as male. In terms of race, 80% identified as Caucasian, 7% as African American, 0% as Asian, 0% as Pacific Islander, and less than 1% as American Indian or Native Alaskan, with the remainder selecting “other” or “Do not know/Unsure”. Nearly 27% of the total population self-identified as Hispanic, whereas the rest identified as non-Hispanic or other. Ninety-five percent of the service members were career firefighters. Participants were classified into three groups: (1) early career defined as less than 10 years in the fire service, (2) mid-career defined as 10–19 years of service, and (3) late career defined as those with 20 years or more of service.2 frequencies. Functional hearing assessment information was obtained for 176 firefighters. Twelve ears could not be assessed due to the presence of occluding cerumen/debris or the visualized perforation of the tympanic membrane, totaling 340 ears for inclusion. Those firefighters with audiometric thresholds exceeding 25 dB for at least one frequency were considered to have impaired hearing. Normal hearing function was indicated in 81% (273/336) of ears in career firefighters by standard audiograms. Conversely, 64% (217/340) of firefighter ears had absent DPOAEs at two or more fF = 12.03, p < 0.001, ηp2 = 0.069), with DPOAE amplitude decreasing with increasing years of service (p < 0.001) and a mean difference of 5.4 dB SPL between the early-career and the late-career firefighters (p < 0.001). Mean DPOAE amplitude decreased by 2.5 dB SPL in late-career firefighters compared to mid-career firefighters (p < 0.05). We also observed that the total number of frequencies where DPOAE responses were absent increased with increasing years in the fire service  = 6.97, p < 0.01).Analyses revealed the significant effect of years in the fire service . While the study population ranged in age from 21 to 59 years, the vast majority (78%) of the population was under 45 years of age. Thus, age was likely not the main factor in the DPOAE findings for this cohort [F = 6.06, p < 0.01, ηp2 = 0.036), but no main effect of age  = 1.39, p = 0.25, ηp2 = 0.013), and no interaction of years of service by age  = 6.06, p = 0.05, ηp2 = 0.024). Furthermore, mean DPOAE amplitude did not vary significantly by age for those with less than 10 years of service. Specifically, firefighters in the highest age brackets (40–49 years and 50+ years) with 10 or more years of service had significantly reduced mean DPOAE amplitudes compared to their age-matched firefighter cohorts with <10 years of service .When looking at age as a factor, correlation analysis revealed that age was significantly associated with reduction in DPOAE amplitude for the frequency range of 5–10 kHz, although this only accounted for a small portion of the variance  = 5.52, p < 0.05, R2 = 0.017, whereas for the higher frequencies the F = 52.50, p < 0.001, R2 = 0.139. Even with a gradual slope, age significantly predicted DPOAE amplitude at both the lower frequency range of 1.4–4 kHz  and the higher frequency range of 5–10 kHz , indicating that for every year increase in age, DPOAE amplitude reduced by −0.113 at the lower frequencies and −0.294 at the higher frequencies.In total, 6.3% of firefighters who participated in the study reported a prior diagnosis of hearing impairment. Overall, 55% of firefighters reported experiencing some combination of hearing symptoms during their career in the fire service. When assessing hearing symptoms experienced following exposure to hazardous noise , the senSpearman’s rank correlation was performed to explore the potential relationships between self-reported hearing symptoms and DPOAE. No significant correlations were observed for the presence of ringing or buzzing in the or for auditory symptoms experienced after noise exposure with self-reported DPOAE . Bonferrt-test was conducted to compare DPOAE amplitude in firefighters who had no concerns about their hearing to firefighters who self-reported concerns about hearing loss. Results found that firefighters who had concerns about hearing loss had lower DPOAE amplitudes  than firefighters who did not have concerns about their hearing . Although imbalance is a less recognized consequence of noise exposure [To explore the relationship between self-reported concerns about hearing and DPOAE amplitude, responses were then converted to binary variables and an independent-samples exposure , firefigNoise exposure is a recognized modifiable risk factor contributing to the development of age-related hearing loss ,6,7,8. INoise is ubiquitous in the fire service and is a leading cause of acquired NIHL in this group, particularly over the span of a long career ,41. The There is mounting evidence that such repeated or prolonged exposure to noise can lead to inner ear structural changes in the presence of normal hearing. This “hidden hearing loss” precedes detectable hearing loss . Our datp < 0.01). Perceived changes in hearing sensitivity were not found to be correlated with DPOAE amplitude. However, the lack of relationship may be related to poor recall or the under-recognition of auditory symptoms as negative health consequences, particularly when they are subtle or less overt. Presumed healthy individuals may be more likely to dismiss symptoms or interpret them as insignificant, which in turn may influence their recall. In general, noise is under-recognized as a potential health hazard [Noise injury is also associated with temporary shifts in hearing sensitivity. Repeated overexposures lead to a permanent change in hearing over time, making temporary changes in hearing after exposure a reliable indicator of noise-induced damage to the ear . Temporah hazard . It is pIn general, firefighters are highly aware of the common symptomology characterizing noise injury, with 55% reporting experiencing at least one auditory symptom following hazardous noise exposure. However, they seemingly do not perceive these symptoms as an actual health hazard, based on the small proportion reporting concerns about their hearing (16%). It is also possible that the determination of “normal hearing” from audiometric findings leads firefighters to under-recognize signs of noise injury. Here, 48% of firefighters reported tinnitus, yet less than 20% had measurable hearing loss on audiometry.Interestingly, firefighters were more concerned about balance than hearing, despite not recognizing imbalance as a potential consequence of hazardous noise exposure . This maAs the aging ear is also subject to changes to the sensory structures of the inner ear, such as OHC loss and atrophy of the stria vascularis ,48,52,53Noise exposure is a modifiable risk factor for hearing loss. Evidence shows that the successful management of modifiable risk factors can be effective in both delaying or preventing disease and reducing healthcare costs . Our resIt is well established that hazardous noise exposure leads to an acceleration of age-related hearing loss and that noise injury leads to the inner ear being more susceptible to early aging ,70,71. TA primary limitation of our study is the lack of an age-matched control group. There are also no reliable comparison data available for accumulated noise exposure over the years. We also acknowledge the potential effects of ototoxic agents on hearing loss in the fire service. However, we note that our focus was on examining the effects of years of service, which serves as a proxy for total noise exposure, on absolute DPOAE amplitude. We also note the inherent challenges of conducting a field study in a large cohort of occupational workers where time constraints prevent the implementation of more comprehensive test batteries. While we controlled the environmental factors that can adversely impact the measurements, challenges remain in implementing these sensitive measures as surveillance tools in the field with potential impact on test–retest reliability.The characterization of DPOAE in a group of relatively young firefighters suggests that firefighters are at risk of NIHL due to their repeated exposure to hazardous noise during their service.Findings indicate that noise injury occurs early and is not detected by standard surveillance methods.The significant associations between subclinical hearing deficits and number of years in service highlight the importance of early detection measures and the need to address the role of a critical modifiable risk factor, noise overexposure, in mitigating hearing loss and reducing the prevalence of cognitive impairment in aging adults.


NDP (norrin cystine knot growth factor NDP) gene. In addition to typical severe ophthalmologic and audiologic defects, the patients showed new pathological features of endolymphatic hydrops (EH), and they also showed acoustic nerves abnormal as described in a very recent report. PCR methods were developed to analyze and diagnose the variation of the family members. This study expands the understanding of the clinical manifestation and pathogenesis of ND and provides a new idea for the treatment of patients in this family and a convenient method for the genetic screen for this ND family.Norrie disease , a rare X-linked recessive genetic disorder, is characterized by congenital blindness and occasionally, sensorineural hearing loss, and developmental delay. The congenital blindness of ND patients is almost untreatable; thus, hearing is particularly important for them. However, the mechanism of hearing loss of ND patients is unclear, and no good treatment is available except wearing hearing-aid. Therefore, revealing the mechanism of hearing loss in ND patients and exploring effective treatment methods are greatly important. In addition, as a serious monogenic genetic disease, convenient gene identification method is important for ND patients and their family members, as well as prenatal diagnosis and preimplantation genetic diagnosis to block intergenerational transmission of pathogenic genes. In this study, a Norrie family with two male patients was reported. This pedigree was ND caused by large fragment deletion of  Norrie disease (ND) is a rare X-chromosome-linked recessive genetic disease, mainly characterized by congenital blindness. Almost all patients are male, and the incidence rate is approximately 1/100,000 . ND patiNDP (norrin cystine knot growth factor NDP) gene mutation is the only genetic factor of ND , and the number is PRJNA8000002.Genomic DNA was extracted from the peripheral blood by a blood genome extraction kit . WES was performed at the Beijing Mykino Medical Laboratory , and the exons were captured using Agilent SureSelect Human All Exon V6 kit, and high-throughput sequencing was performed using Illumina HiSeq X-10 sequencer. Sequence comparison and splicing used software, such as BWA 26 (Burrow-Wheeler Aligner 26), SAMtools27 , and PicardFiltered pathogenic variants of WES were validated by Sanger sequencing. The PCR primers were designed by Primer 5 , and theThe propositus (III-1) was a 10-year-old boy, and the pedigree was shown in III-1 had normal ear structure, growth, and behavior and no peripheral vascular disease. Pure tone audiometry showed moderate sensorineural hearing loss in both ears, with evident low frequency hearing loss. The average hearing threshold of speech frequency  was 55 dB, and the high-frequency  hearing loss was mild; the hearing threshold map showed an upward pattern . The wavPatient II-1 was 39 years old. According to the family description, the patient had normal hearing when he was young, and hearing loss occurred when he was approximately 20 years old; the condition gradually worsened, and he had difficulty in communicating with words. He had normal ear structure, normal growth, and development, and not peripheral vascular disease. Due to the patient’s difficulty in communication, pure tone audiometry could not be performed, and air-conducting m-ASSR examination showed severe sensorineural hearing loss at almost all the frequencies in both ears. At 500 Hz, the response threshold of the right ear was 75 dB, and that of the left ear was only 35 dB. In the frequency region of 1,000–4,000 Hz, the response threshold of binaural exceeded 75 dB except for the 65 dB of the left ear at 2,000 Hz . ABR exaThe cognitive function of the patients was evaluated by MMSE and MOCA scales, and the spatial orientation, long-term memory, and computational ability decreased in III-1; the visual–spatial and logical ability tests could not be completed due to visual problems. II-1 could not be evaluated by scales due to difficulty in communication. The craniocerebral MRI results of II-1 showed few ischemic lesions in the brain, and the structure of hippocampus and fronto temporal cortex was normal without obvious atrophy . HoweverNDP gene located on the X chromosome , and then developed into severe, symmetric, flat loss at approximately 35 years old. However, after severe hearing loss from approximately 35 to nearly 60 years old, the progress became stable . SimilarNDP mutation alone does not cause EH. Another possibility is that EH is an uncommon phenotype of ND, which was missed due to insufficient attention in previous studies. We hope that future researchers can focus on this phenotype and further confirm this symptom. In summary, the symptom of EH in ND patients were reported for the first time; it may be one of the mechanisms of hearing loss of ND patients. These discoveries expanded the understanding of ND, and provided a new idea for the treatment of ND patients.Endolymph is closely related to vestibular and auditory sensors, and its production and absorption reflux is a circulatory process. If the reflux is blocked or absorption is impaired, then it can lead to excessive endolymphatic retention known as EH. EH can cause tinnitus, deafness, and vertigo in vestibular and auditory organs. Distension of the membranes in EH starts within the apex, and then the accumulated endolymph extends to the middle and base turn because the basilar membrane is wider and softer in the apex than that in the base of the cochlea . Therefo± mice, a model of EH, the loss of helical ganglion cells shows a topographical pattern ; this pattern mirrors the progression of EH. Interestingly, precedes the eventual loss of ganglion cells, the expression of apoptosis markers, such as caspase 3, 8, and 9, increased in spiral ganglion cells correspondingly  and aging-related hearing loss (lost vascular density). The study on ND knockout mice found that the stria vascularis was the most affected structure. With the increase of age, disorganization and enlarged vessels were observed in the stria vascularis, as was the lack of a well-developed capillary bed and eventual loss of two-thirds of the vessels in the stria vascularis. A gradual loss of outer hair cells was observed; it is particularly consistent with auditory threshold increases. As the hearing loss progressed, the spiral ganglion began to degenerate. These lesions were particularly pronounced in the apical portion of the cochlea . So degeNDP mutation. Wnt signaling pathway plays a key role in the formation of blood vessels in various organs, including the eye. Various Wnt ligands are widely distributed in the retina and inner ear of developing mice  is often used. However, when the deletion fragments have been identified, these methods are expensive and unnecessary. As for the reported Q-PCR methods  can be found below: NCBI, SRA The studies involving human participants were reviewed and approved by the Ethics Committee of the Binzhou Medical University Hospital. Written informed consent to participate in this study was provided by the participants’ legal guardian/next of kin.YG, ZL, and XZ performed the research and wrote the manuscript. JS and WL contributed to clinical examination. QX contributed to patient recruitment and data collection. SS and HZ analyzed the data. YW and QZ revised the manuscript. JC and XL contributed to project design and revised the manuscript. 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.


Criteria for identifying and quantifying hearing loss have been devised.Cisplatin is used frequently as an antineoplastic drug in the treatment of many different cancers. However, when used in doses over 360mg/mTo describe the features of different hearing loss classification systems and to identify their implications and use in oncologic patients.Hearing loss was classified in 31 patients before and after chemotherapy, according to different criteria, assessing the sensitivity and specificity of each classification system.Hearing loss results were highly variable (ranging from 29% to 61%). Only 4 of 31 subjects with post-therapy hearing loss were identified by all the methods. A few subjects with hearing loss were classified as normal hearing in some of the criteria. A normal PTA was found in 18 of 31 subjects in the post-treatment evaluation.None of the criteria assesses the complaints of patients. The criteria described in this study were inadequate to identify hearing loss following chemotherapy, requiring additional information for physicians to better understand the hearing losses and their implications for the quality of life of patients. Side effects include: nausea, vomiting, myelosuppression nephrotoxicity, central and peripheral neuropathies, and ototoxicity . All patients were consulted about their participation in the study, and signed a free informed consent form. Audiological assessments were carried out in 31 subjects seen at the Audiology unit; inclusion criteria were as follows: cisplatin chemotherapy only; absence of ontological complaints; no radiotherapy in the head and neck, and pre– and post-therapy audiological assessments.There were 16 male and 15 female patients. Age ranged from 7 to 66 years; the mean age was 28 years.All subjects in this study underwent full audiological testing before, during and after chemotherapy with CDDP; however, only conventional pure tone audiometry data were computed. A Madsen, Orbiter model version 922 audiometer was used for measuring air and bone conduction pure tone auditory thresholds (in dBHL) . Fisher's exact test was applied to study the significance of each method. The GraphPad Prism version 2.0 software was used. The significance level was < 0.05.Davis and Silverman'sStatistical analysis consisted of specificity and sensitivity calculations for each criterion; Fisher's test was applied to calculate the p value, as shown on the Table below.•Only 4 subjects with post-therapy hearing loss were identified by all criteria; 7 subjects had no hearing loss in any of the criteria applied in this study.•Thirteen of 18 subjects with normal pure tone audiometry (according to Davis and SilvermanThe distribution of audiometric thresholds in the study sample showed that:Brock et al.'sAuditory losses due to cisplatin ototoxicity are generally symmetrical, bilateral, initially affecting high frequencies, followed by middle and low frequencies (Testa et al.Changes were not always detected at the beginning by the criteria described above. A major point about these criteria is that they do not take the complaints of patients into account, which would be extremely important. Additionally, the impact on a patient's life may not be proportional to the degree of hearing loss, since this impact depends on factors such as social and professional activities and personal aspects. The growing concern on the part of oncologists should lead to careful prevention of hearing loss, comprising periodic testing and close monitoring of hearing losses, not waiting for patients to complain before becoming concerned with hearing; this means preventing the onset of hearing loss and not only rehabilitation when hearing cannot be recovered any longer.A relevant point was that NCIAudiologically, there are monitoring proposals using conventional frequencies (Testa et al.A further discrepancy is the grade II in this criterion. This grade includes patients that had a 25 to 90 dB elevation of the auditory threshold. A 25 dB threshold elevation may go unnoticed or may cause minimal difficulty; it may even be classified as mild hearing loss (Davis and SilvermanThe 40 dB reference in Brock et al.'sThe ASHADavis and SilvermanThe same subject with hearing loss was not always classified by all of the criteria above; thus, the sensitivity and/or specificity of each criterion needs to be known, as small changes in hearing are not always detected by these instruments. Important hearing losses are easily detected by any of these tools; however, a classification instrument should detect small changes so that oncologists and speech therapists may carefully monitor the hearing function of these patients, to avoid major loss of function.The classification of hearing loss in audiological evaluations during oncological therapy aims firstly to identify ototoxic effects, especially at high frequencies. This makes it possible for physicians to be alert and change therapy protocol measures. Secondly, the classification should indicate at which point patients will suffer the social, educational and professional implications of hearing loss. Impact differs in adults and children, both in terms of the degree of hearing loss and the affected frequencies. A classification should be able to show the progression of hearing loss.Knight et al.LibermanMarini et al.Teles et al.Gupta et al.According to the literature we cited, changes in hearing were not always detected at the beginning in oncological patients by the criteria described in this study. The main criticism of these criteria is that they do not include patient complaints, essential for understanding the impact of hearing loss on patient's lives, which are not always proportional do the degree of loss, but depend on factors such as personal, social and professional activities. The growing concern of oncologists should lead to preventive care to avoid hearing loss; this includes periodic testing and close monitoring of auditory changes before patients complain to avoid hearing loss rather than just rehabilitation when reversion is no longer possible.Each classification tool should be assessed according to its purpose. In the case of hearing loss associated with oncological therapy, we believe it is relevant not only to identify, classify and quantify auditory losses, but also to investigate eventual complains and the impact that hearing loss may cause on the quality of life of patients. It is impossible to define the degree of hearing loss that results in complaints and difficulties for patients using only the criteria mentioned above for classifying auditory losses. Thus, identifying and validating the complaints of patients is just as important as audiological evaluations and classification of losses according to charts.It is extremely important to note that the criterion used for classifying hearing losses in oncological patients should be able to identify the beginning of loss, thus avoiding unnecessary adverse effects and preventing auditory losses due to oncological therapy.We found that Davis and Silverman'sHowever, all of these criteria underestimated the description of identified auditory alterations; additional information was required to help physicians to understand the true implications of hearing loss in each case.A common code is needed between audiologists and oncologists to increase our understanding and improve the therapy of these patients; not only should the grade and type of hearing loss be described, but also the impact on the lives of patients.


Mycoplasma hyopneumoniae. One vaccine contained PCV2a and the other contained PCV2b, and both were administered on a farm suffering from subclinical PCV2d infection and enzootic pneumonia. A total of 180 pigs were randomly divided into 3 groups . Bivalent vaccination significantly improved growth performance in both vaccinated groups as compared to the unvaccinated (UnVac) group. Growth performance measured by body weight and average daily weight gain (ADWG) was not significantly different between the two bivalent-vaccinated groups (VacA and VacB). Both bivalent vaccines elicited high levels of neutralizing antibodies and interferon-γ secreting cells (IFN-γ-SC) against PCV2d, leading to a reduction in the levels of PCV2d blood viral load as compared to unvaccinated animals. Similarly, both bivalent vaccines elicited high levels of IFN-γ-SC against M. hyopneumoniae that reduced the level of M. hyopneumoniae laryngeal viral loads as compared to unvaccinated animals. Significant differences in severity of lung and lymphoid lesions were observed in both vaccinated groups as compared to the UnVac group. These comparative field data demonstrated that both bivalent vaccines are good candidates for controlling subclinical PCV2d infection and enzootic pneumonia in swine farms suffering from an existing infection.The objective of this study was to compare two different bivalent vaccines containing porcine circovirus type 2 (PCV2) and  Mycoplasma hyopneumoniae is recognized as the primary causative agent of the so-called enzootic pneumonia. The disease causes significant economic loss, also due to growth retardation and the increased cost of antimicrobial medication (Porcine circovirus type 2 (PCV2) is the main etiological agent of porcine circovirus-associated disease (PCVAD) , 2. Sincdication , 7.M. hyopneumoniae is a better choice in the control of PRDC in herds suffering from severe respiratory disease.Complications from both subclinical PCV2 infection and enzootic pneumonia were involved in the porcine respiratory disease complex (PRDC). PRDC devastates swine herds through decreased growth rate and poor feed efficiency results in an extended time to market , 9. VaccM. hyopneumoniae are commercially available  may offer better protection than a heterologous (non-matched genotype) vaccination and challenge for PCV2 . Howevervailable , 19. Forvailable . To dateAll of the methods were approved by the Seoul National University Institutional Animal Care and Use and Ethics Committee (SNU-210518-3).M. hyopneumoniae. The farm was selected on the basis of subclinical PCV2 infection and enzootic pneumonia. The farm consistently had respiratory problems due to poor growth in the late post-weaning and growing stages. Clinical signs first appeared at approximately 8–11 weeks of age and reached peak mortality (~3–5%) between 10 and 15 weeks of age. PCV2d was detected in serum from 3 pigs with poor growth, where log10 DNA copies/ml ranged from 2.46 to 3.34. These values were consistent with the definition of subclinical PCV2 infection  was maintained with the absence of active porcine reproductive and respiratory syndrome virus (PRRSV) circulation . Sows were not previously immunized against PCV2 and nfection , 22. ThrTo reduce sow variation, six pigs (21-day-old) per sow were randomly selected by the random number generator function  and evenly allocated to each of the three groups. A total of 180 pigs were randomly divided into 3 groups  within the same software and function .M. hyopneumoniae  at the right side of the neck in accordance with the manufacturer's directions. According to the manufacturer's instruction, pigs in the VacB group were intramuscularly vaccinated with a 1.0 ml dose of the bivalent vaccine containing PCV2b and M. hyopneumoniae  in the same anatomical location. Pigs in the unvaccinated (UnVac) group were injected with 2.0 ml of phosphate-buffered saline  in the same anatomical location. Sample collection of blood and laryngeal swabs were performed at 0 (21 days old), 28 (49 days old), 49 (70 days old), and 91 (112 days old) dpv.At 0 day post-vaccination , pigs in the VacA group were intramuscularly vaccinated with a 2.0 ml dose of the bivalent vaccine containing PCV2a and The pigs were daily monitored for abnormal clinical signs and weekly scored using scores ranging from 0  to 6  . The morThe live weight of each pig was measured at 21 (0 dpv), 70 (49 dpv), and 175 (154 dpv) days of age. Over two time periods, (i) between 21 and 70 days old and (ii) between 70 and 175 days old, the ADWG (grams/pig/day) was analyzed. During the different production stages, ADWG was calculated as the difference between the starting and final weight divided by the duration of the stage. Data for culled or dead pigs were also included in the calculation.TM-CAT CAT GTC CAC ATT CCA G-3′ Black Hole Quencher) were designed for detecting specific Capsid-coding region of PCV2d only . An extracted DNA sample was used for the quantification of PCV2d genomic DNA copy numbers by real-time PCR . The forV2d only .M. hyopneumoniae genomic DNA copy numbers by real-time PCR . An extracted DNA sample was used for the quantification of time PCR . The foreumoniae .M. hyopneumoniae  and PCV2 . According to the instruction of the manufacturer for each kit, serum samples were recorded as positive for M. hyopneumoniae antibody if the S/P ratio (sample-to-positive ratio) was ≥0.4. For anti-PCV2 antibodies, samples were recorded as positive if the reciprocal ELISA titer was >350. A serum virus neutralization test against PCV2d was used to test the serum sample (Enzyme-linked immunosorbent assay (ELISA) was performed to measure antibodies against m sample –28.M. hyopneumoniae-specific interferon-γ secreting cells (IFN-γ-SC) in peripheral blood mononuclear cells (PBMCs) was measured by ELISPOT assay  1640 medium supplemented with 10% fetal bovine serum  were seeded onto plates and precoated overnight with anti-porcine IFN-γ monoclonal antibody  and incubated with PCV2d (20 mg/ml), M. hyopneumopniaee (4 mg/ml), phytohemagglutinin  as a positive control, or PBS as a negative control for 20 h at 37°C in a 5% humidified CO2 atmosphere. The wells were washed five times with PBS (200 ml per well) and thereafter, the procedure followed instructions of the manufacturer using a commercial ELISpot assay kit (Mabtech). The spots on the membranes were read by an automated ELISpot reader . The results were expressed as the number of responding cells/million PBMC.The number of PCV2d-specific and OT assay , 29. BriTo estimate the percentage of the lung affected by pneumonia, two pathologists (Chae and one graduate student) at the Seoul National University  scored the severity of macroscopic lung lesions as previously described . The col10 and log2 values, prior to statistical analysis. The collected data were assessed by the Shapiro-Wilk test for a normal distribution. Then, one-way ANOVA was performed to determine if there were statistically significant differences between different groups at each time point. For further evaluation, a post-hoc test for a pairwise comparison with Tukey's adjustment was conducted with a statistical significance result from a one-way ANOVA test. In case of the normality, assumption was not met and the Kruskal-Wallis test was performed. Results, which showed a statistical significance from the Kruskal-Wallis test, were further evaluated with the Mann-Whitney test to include Tukey's adjustment to compare the differences among the groups. The value of p < 0.05 was considered significant and reported in p-values.Real-time PCR and neutralizing antibody data were recalculated to logp < 0.05) lower respiratory signs than those of unvaccinated animals from the UnVacA group at 14–112 dpv. There were no differences in respiratory signs between the two vaccinated groups (VacA and VacB).Two vaccinated animals from the VacA and VacB groups had significantly (p < 0.05) higher than that of the unvaccinated group (UnVac). Overall (3–175 days of age), the ADWG of the vaccinated groups (VacA and VacB) was significantly (p < 0.05) higher than that of the unvaccinated group (UnVac) , the ADWG of the vaccinated groups (VacA and VacB) was significantly ( (UnVac) .M. hyopneumoniae and PCV2d that was detected with PCR testing, and Pasteurella multocida was isolated from the lungs at 77 and 85 days of age. One pig from the VacB group died of pleuropneumonia caused by coinfection with Actinobacillus pleuropneumoniae and Glaesserella parasuis that was determined through isolation from the lungs at 81 days of age. A total of 4 pigs were died in the UnVac group; two pigs were died of bronchopneumonia as determined by a combination of M. hyopneumoniae that was detected with PCR, and P. multocida and Trueperella pyogenes that were isolated from the lungs at 69 and 85 days of age. The other two pigs were died from bronchopneumonia as determined by a combination of M. hyopneumoniae and PCV2d that were detected with PCR, and G. parasuis that was isolated from the lungs at 81 and 86 days of age.A total of 2 pigs died in the VacA group of severe pneumonia as determined by a combination of p < 0.05) lower than that of unvaccinated groups (UnVac) at 28, 49, and 91 dpv (The PCV2 DNA blood loads from vaccinated groups (VacA and VacB) were significantly (d 91 dpv . Two of M. hyopneumoniae DNA loads in the larynx was significantly (p < 0.05) lower in the vaccinated groups (VacA and VacB) than in those of the unvaccinated group (UnVac) between 28 and 91 dpv (M. hyopneumoniae DNA loads in their larynx.The amount of d 91 dpv . Throughp < 0.05) higher in their PCV2 ELISA titers (The vaccinated groups (VacA and VacB) were measured significantly  higher in their M. hyopneumoniae ELISA S/P ratios (M. hyopneumoniae ELISA S/P ratios or IFN-γ-SC were observed in the two vaccinated (VacA and VacB) groups throughout the entire field trial.Both vaccinated groups (VacA and VacB) were measured significantly  effectively reduced macroscopic lung lesion scores, microscopic lung, lymphoid lesion scores, and the numbers of lymphoid PCV2-positive cells when compared to the unvaccinated group (UnVac) at 154 dpv . ThroughM. hyopneumoniae provide equal protection for pigs in herds suffering from subclinical PCV2d infection and enzootic pneumonia. The common denominator of PCV2d and M. hyopneumoniae infection is weight loss, so it was important to evaluate the improvement of growth performance in the comparative field trial. Vaccination of pigs with both evaluated bivalent vaccines (two groups) resulted in a significantly improved growth performance as compared to the pigs in the unvaccinated group. Significant differences in growth performance as measured by body weight and ADWG were not found between the two bivalent-vaccinated groups.The results of this comparative field trial demonstrate that PCV2a- and PCV2b-based bivalent vaccines that also contain M. hyopneumoniae combination vaccines induced protective immunity by reducing PCV2 blood viral load and M. hyopneumoniae laryngeal load while simultaneously reducing lung and lymphoid lesions, thereby controlling these two diseases  can be found in the article/supplementary material.The animal study was reviewed and approved by Seoul National University Institutional Animal Care and Use Committee (SNU-210518-3).HC and TO contributed to the performance of the experimental trials, data analysis, and writing of the manuscript. HC, TO, and JS contributed to the preparation of the inoculum and lab analysis. CC contributed to the development of the protocol, design of the study, review of the final manuscript, and the approval for publication. All authors read and approved the final manuscript.The author's research was supported by contract research funds (Grant No. 550-20190068) of the Research Institute for Veterinary Science (RIVS) from the College of Veterinary Medicine and by the BK 21 Plus Program (Grant No. 5260-20150100) for Creative Veterinary Science Research.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. 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RNA-binding proteins (RBPs) and circular RNAs (circRNAs) play important roles in glioblastoma multiforme (GBM). Aerobic glycolysis is a metabolic characteristic of GBM. However, the roles of RBPs and circRNAs in aerobic glycolysis in GBM remain unclear. The aim of this study is to explore the mechanisms by which RBPs and circRNAs regulate aerobic glycolysis in GBM cells.RNA sequencing and circRNA microarray analysis were performed to identify RBPs and circRNAs for further study. Mass spectrometry validated the encoded protein and its interacting proteins. Quantitative reverse transcription PCR and western blot assays were used to determine the mRNA and protein expression, respectively. Furthermore, immunofluorescence and fluorescence in situ hybridization assays were used to determine the protein and RNA localization, respectively. Glucose and lactate measurement assays, Seahorse XF glycolysis stress assays and cell viability assays were conducted to investigate the effects on glycolysis and proliferation in GBM cells.We selected zinc finger CCHC-type and RNA-binding motif 1 (ZCRB1) and circRNA HEAT repeat containing 5B (circHEATR5B) as candidates for this study. These genes were expressed at low levels in GBM tissues and cells. Both ZCRB1 and circHEATR5B overexpression suppressed aerobic glycolysis and proliferation in GBM cells. ZCRB1 overexpression promoted the Alu element-mediated formation of circHEATR5B. In addition, circHEATR5B encoded a novel protein HEATR5B-881aa which interacted directly with Jumonji C-domain-containing 5 (JMJD5) and reduced its stability by phosphorylating S361. JMJD5 knockdown increased pyruvate kinase M2 (PKM2) enzymatic activity and suppressed glycolysis and proliferation in GBM cells. Finally, ZCRB1, circHEATR5B and HEATR5B-881aa overexpression inhibited GBM xenograft growth and prolonged the survival time of nude mice.This study reveals a novel mechanism of regulating aerobic glycolysis and proliferation in GBM cells through the ZCRB1/circHEATR5B/HEATR5B-881aa/JMJD5/PKM2 pathway, which can provide novel strategies and potential targets for GBM therapy.The online version contains supplementary material available at 10.1186/s13046-022-02374-6. Glioblastoma multiforme (GBM) is a primary malignant tumor in the central nervous system with high morbidity and mortality rates, characterized by aggressive growth and a high recurrence rate . The mosMost tumor cells prefer to obtain energy through glycolysis rather than oxidative phosphorylation even in the presence of sufficient oxygen, a phenomenon known as aerobic glycolysis or the Warburg effect,  as it isAs one of the three rate-limiting enzymes of glycolysis, pyruvate kinase includes four isozymes, of which M2-type pyruvate kinase (PKM2) is mainly expressed in rapidly proliferating cells, especially tumor cells, and it plays a crucial role in the development of tumors, such as GBM,  by regulZCRB1 gene, mapped to chromosome 12q12. Researchers have demonstrated that ZCRB1 can bind specifically to the 5′-UTR of SARS-CoV RNA and promote viral RNA synthesis  ATP  for 30–60 min at 30 °C. The kinase reaction was terminated by adding 12.5 μl 5 × SDS-PAGE loading buffer and boiling for 10 min. Then, the samples were resolved by SDS-PAGE gel electrophoresis. Radioactive signals were detected by autoradiography through a phosphor screen. The gels were subjected to Coomassie brilliant blue staining and destaining to visualize the protein bands.In brief, 10 μg purified GST, GST-JMJD5(-WT), or GST-JMJD5-S361A proteins were incubated with 5 μg recombinant active HEATR5B-881aa in 50 μl kinase buffer  containing 5 μCi [γ-SDS-PAGE gel electrophoresis separated proteins and the targeted protein bands were excised from the gel. After elution, reduction and alkylation, the proteins were digested at 37 °C overnight. Peptides were collected, desalted and analyzed by timsTOF pro mass spectrometer . Sequence and site identification were analyzed using NCBI nonredundant protein database with Mascot Daemon.Cycloheximide, a protein synthesis inhibitor, was used to determine JMJD5 half-life. U251 cells transfected with JMJD5-WT, JMJD5-S361A, or JMJD5-S361E were treated with 100 mg/ml cycloheximide  and collected in 0, 2, 4, 8, 10 h. Then, total proteins were prepared and detected by western blot assays.5 cells were subcutaneously injected into the right flank of each nude mouse. Tumor volumes were measured every 4 days and calculated by formula: volume (mm3) = length×width2/2. The mice were sacrificed and then xenograft tumors were separated at the endpoints. For orthotopic xenografts, nude mice were injected with 3 × 105 cells stereoscopically into the right striatum. The surviving numbers of nude mice were recorded, and Kaplan-Meier curves were used for survival analysis.Four-week-old nude mice (BALB/c) were purchased from Beijing Vital River Laboratory for in vivo study. The experiments were conducted strictly following the protocols approved by the Ethics Committee of China Medical University. The stably transfected U251 and U373 cells were selected and divided into five groups. For subcutaneous xenografts, 3 × 10P-value< 0.05.All experimental data were indicated as mean ± SD. GraphPad Prism 5.01 was used for statistical analysis. Comparison between groups was analyzed by Student’s t-test, one-way ANOVA, or two-way ANOVA. Statistical significance was determined by In our study, differentially expressed genes between GBM and NBTs were screened by RNA-seq  and two internal ribosome entry sites (IRESs) and might encode an 881 amino acid protein (Additional file In U251 and U373 cells, we found that HEATR5B-881aa decreased with circHEATR5B knockdown and increased with circHEATR5B overexpression Fig. F. We theTo investigate the biological functions of HEATR5B-881aa, we upregulated HEATR5B-881aa using OV-HEATR5B-881aa plasmids and downregulated HEATR5B-881aa using sh-circHEATR5B plasmids without affecting HEATR5B protein in U251 and U373 cells (Fig. To further exclude the possibility that circHEATR5B, but not HEATR5B-881aa, induced the above effects, we mutated the ATG start codon of circHEATR5B (Fig. IF assays revealed that HEATR5B-881aa was mainly localized in the cytoplasm Fig. A. To revIn GBM tissues and cells, JMJD5 was significantly elevated and positively correlated with the pathological grades of glioma Fig. B and C. Then, we found that JMJD5 protein decreased as HEATR5B-881aa increased and that it increased as HEATR5B-881aa decreased Fig. I. HoweveTo further investigate the mechanisms by which HEATR5B-881aa downregulated JMJD5 expression, IF assays were performed and showed that HEATR5B-881aa colocalized with JMJD5 in the cytoplasm of U251 and U373 cells (Fig. In vitro kinase assays revealed that HEATR5B-881aa phosphorylated JMJD5 protein Fig. E. By anaSubsequently, we transfected JMJD5-WT, JMJD5-S361A, and JMJD5-S361E plasmids into U251 and U373 cells and found that JMJD5 with low phosphorylation levels significantly increased glucose consumption, lactate production, proliferation, and ECAR in U251 and U373 cells and promoted xenograft tumor growth, while JMJD5 with high phosphorylation levels significantly inhibited these effects (Fig. Finally, nude mouse xenograft models were used to clarify the in vivo tumor-suppressive effects of ZCRB1, circHEATR5B, and HEATR5B-881aa. The grouping is shown in Fig. Survival analysis of nude mice with orthotopic xenograft tumors revealed that the groups of ZCRB1, circHEATR5B, and HEATR5B-881aa overexpression had longer survival time than the negative control group, and the three-combined group had the longest survival time (Fig. RBPs have been reported to play an important regulatory role in tumor glycolysis . TherefoPrevious studies have indicated that circRNAs exert essential functions in GBM development –40. To iWe found that ZCRB1 overexpression increased nascent circHEATR5B expression without changing its half-life, indicating that ZCRB1 upregulates circHEATR5B by promoting its formation rather than inhibiting its degradation. To elucidate the mechanisms by which ZCRB1 promoted circHEATR5B formation, we first predicted the binding of ZCRB1 and circHEATR5B flanking sequences by RBPmap database and further confirmed it experimentally. It has been reported that RBPs such as QKI and Mbl can promote circRNA formation by binding flanking sequences , 43. BacPrevious studies have proven that circRNAs have various biological functions, such as acting as microRNA or RBP sponges, regulating gene splicing and transcription, and translation into proteins . Due to Studies have shown that proteins encoded by circRNAs can exert biological functions in tumor development , 48, 49.+ concentration measured by the Seahorse apparatus.To investigate the possible mechanisms by which HEATR5B-881aa regulated glycolysis and proliferation in GBM cells, we performed Co-IP assays coupled with mass spectrometry and selected JMJD5, which has been reported in breast cancer to bind and induce PKM2 dimerization, thereby regulating glucose metabolism . PKM2 diWe found that HEATR5B-881aa reduced JMJD5 protein expression without influencing JMJD5 mRNA expression, and JMJD5 overexpression rescued glycolytic and proliferative inhibition induced by HEATR5B-881aa, indicating that HEATR5B-881aa inhibits glycolysis and proliferation in GBM cells by downregulating JMJD5 at the protein level. However, the mechanisms by which HEATR5B-881aa decreased JMJD5 protein levels remained unclear. We first identified a direct interaction between HEATR5B-881aa and JMJD5. Through HEATR5B-881aa domain analysis, we speculated that HEATR5B-881aa might regulate JMJD5 by phosphorylation modification. Further studies revealed that HEATR5B-881aa phosphorylated JMJD5 on S361 in vitro and in vivo. In addition, we found that JMJD5 expression was reduced as the S361 phosphorylation level increased, indicating that HEATR5B-881aa reduces JMJD5 by S361 phosphorylation. It has been reported that phosphorylation can affect protein stability . In thisFinally, in vivo experiments showed that ZCRB1, circHEATR5B, or HEATR5B-881aa overexpression could inhibit the growth of subcutaneous GBM xenograft tumors and prolong the survival time of nude mice with orthotopic GBM xenograft tumors; furthermore, their combination had the strongest tumor-suppressive effect. These results further clarify the potential clinical value of ZCRB1, circHEATR5B, and HEATR5B-881aa in GBM.This study demonstrates for the first time that low expression of the RNA-binding protein ZCRB1 inhibits circHEATR5B formation, which further reduces expression of the encoded HEATR5B-881aa, resulting in the reduced phosphorylation and thus increased stability of JMJD5. High expression of JMJD5 induces PKM2 dimerization, thus promoting aerobic glycolysis and proliferation in GBM cells. This study reveals a novel mechanism of regulating aerobic glycolysis and proliferation in GBM cells through the ZCRB1/circHEATR5B/HEATR5B-881aa/JMJD5/PKM2 pathway, which can provide novel strategies and potential targets for GBM therapy.Additional file 1: Figure S1. Screening for ZCRB1, the mRNA expression of ZCRB1, and the transfection efficiency of ZCRB1 plasmids. Figure S2. Screening for circHEATR5B, the mRNA expression of HEATR5B, and the transfection efficiency of circHEATR5B plasmids. Figure S3. The effects of ZCRB1 on circHEATR5B stability and the binding prediction of ZCRB1 and circHEATR5B. Figure S4. The prediction of the potential encoding capacity of circHEATR5B and the expression of HEATR5B protein. Figure S5. Statistical analysis of western blot assays and ECAR parameter calculations. Figure S6. Screening for JMJD5 and the transfection efficiency of JMJD5 plasmids. Figure S7. JMJD5 phosphorylation induced by HEATR5B-881aa downregulated JMJD5 expression, which was reversed by MG132 treatment.Additional file 2: Table S1. The primers for qRT–PCR assays. Table S2. The primers for RIP assays. Table S3. Top ten differentially expressed RBPs in GBM tissues screened by RNA-seq. Table S4. Top ten differentially expressed circRNAs in ZCRB1-upregulated GBM cells screened by circRNA microarray analysis. Table S5. The divergent and convergent primers of circHEATR5B and β-actin.


Several factors have been postulated to elicit the etiology of idiopathic sudden sensorineural hearing loss. Through a bibliographic review, we made a critical analysis of the different etiopathogenic aspects of its clinical manifestation. The most recent studies concerning the possible causes of sudden hearing loss suggest vascular disorders, rupture of the inner ear membrane and autoimmune diseases; however, viral infections have received a great deal of attention in recent years. Little is known about the mechanism of sudden hearing loss. Viruses can cause sudden hearing loss in an acute infection, however the latent form, and its possible reactivation have also been considered as explanations of the cochlear injury mechanism. Even though hearing loss can be explained by a blood viscosity change, experimental and clinical studies do not show any evidence of labyrinthine fibrosis and new bone formation, or labyrinthine membrane breaks. These findings are not in agreement with vascular and rupture membrane factors, respectively. The eventual presence of antibodies against the inner ear suggests that sudden hearing loss pathogenesis may be of autoimmune nature, but the difficulty in establishing the correlation of its morphological and clinical aspects to the hearing loss also do not help to support this statement. Sudden hearing loss is still a controversial and obscure subject in several aspects. Sudden hearing loss (SHL) means a sensorineural hearing loss of sudden onset or one that happens in minutes, hours or even in a few days. The hearing impairment varies as far as intensity and sound frequency are concerned, and in general, it is unilateral (98-99% of the cases)It may be missed in children, specially when it is unilateral. Tinnitus frequently follow the symptoms of idiopathic sudden hearing loss (ISHL) (70% of the cases) and diziness is sometimes present (up to 40% of the cases), thus completing the triad of Ménière. About 10% of dizziness cases may be incapacitating and associated to nausea and vomit. There may also be ear fullness, headache and viral infection symptoms of the upper airwaysOne of the few emergencies in otology, the ISHL affects mostly those in their fourth decade of life, involving both the right and the left ears in equal proportions, and the same trend is seen genderwiseIt affects 5 to 20 persons for each 100 thousand individuals. It is estimated that there are approximately 4 thousand new cases per year in the United States, and 15 thousand all over the worldIn Brazil we lack good epidemiological references as to the true incidence of SHL, due to the difficulty in safely assessing its incidence. Poor cultural and socioeconomical conditions, together with the possibility of spontaneous recovery before seeking medical help or the very disregard towards light symptoms are responsible for this epidemiological pictureAlthough no seasonal influence has been proven for ISHL, it is known that patients with such disorder are mainly affected after an upper airway infection or herpes labial infectionThe hearing loss (HL) caused by ISHL may become permanent or regress either totally or partially. Many authors report that spontaneous recovery happens in 45 to 60% of the patientsSince ISHL physiopathology is still unclear, there are disagreements as to its true causeHaving stated that, our goal is to assess ISHL as to its main and possible etiology and physiopathology.Prosper Ménière was probably the first author to clearly describe a case of SHL without a definitive causeLater, Rasmussen in 1949, apud Fowler (1950) and Hallberg (1956), report cases of ISHL and discuss the disease origin, proposing the vascular origin and the neuritis of the VIII nerve as possible causes. Following that, Lindsay & Zuidema (1950) also studied patients with SHL and discuss possible causes. According to these authors, vascular diseases such as thrombosis could justify the hearing loss, however the age range did not match; hemorrhage would be another possibility, provided the patients had previous history of coagulation problems or head injury. They then consider, although hypothetic, the possibility of vasospasms, besides infection as the possible cause for hearing loss (HL), what could then justify this variable pattern of auditory and vestibular involvement.Back in 1950, Fowler present some SHL cases and stresses that most of the patients had a psychosomatic disorder associated. He reports not only the importance of the supratentorial disorder, but also the change in blood viscosity, as possible causes for SHL. It was also in 1950 that Opheim, apud Hallberg (1956), reported cases of SHL, highlighting the possibility of an acute increase in labyrinth pressure as being responsible for such happening.Later, in 1952, Moulonguet and Bouche, apud Hallberg (1956), reported the case of a young physician with SHL after having used an intravenous medication, and also considered that the arteriolar spasm caused by this drug caused the clinical signs and symptoms in this patient.Disorders of the labyrinthine circulation such as the occlusion of microvessels in the inner ear and cochlear hemorrhage have been considered as possible causes of ISHL, specially because they are of sudden onset, notwithstanding, there are a number of non-compatible aspects, such as age range, risk group and prognosisIt may be that an alteration in blood viscosity caused by a prior disease be consider a predisposing factor, however, not as a primary cause of sudden hearing lossIn 1957, Van Dishoeck & BiermanHeller & LindenbergAutoimmune causes are included as possible causes for ISHL. In 1979, McCabe was the first to associate SHL and progressive sensorineural hearing impairment with autoimmune diseases and to consider it because it can be treated. Not always identified, diseases such as rheumatoid arthritis, lupus, nodose polyarthritis and others are possible mediators of the antibody and immunocomplex processes that may be involved with the origin of the sudden hearing lossPerilymphatic fistulas, in some cases may also explain the hearing loss and its spontaneous recovery in patients with ISHLThis study was carried out with a survey of the different scientific papers related to the theme, with a critical and comparative analysis of these papers.A cause of great expectation, ISHL is a terrible experience for the patient. The sudden silence, followed, sometimes, by tinnitus and/or vertigo, represents, not only the loss of inner ear function, but also the impairment to the patient's psychological status. Although not very relevant, it is an otologic emergency, which bears no established cause or physiopathology and, therefore, without established treatment. Thus, it is a problem that places the physician in a difficult clinical situation. As to clinical evolution and hearing recovery, they are both question marks in the course of the disease.The definition of ISHL is not universal. It is a sensorineural hearing loss that may vary as to the concept of the word “sudden”. There are cases in which the hearing loss may be sudden or happen in a short period o time until it becomes completeThe etiology of the ISHL, as the very name states (idiopathic), is still unknown, what makes the topic still very controversial. In only 10% of the cases the cause can be foundThe emotional factor as possible cause of ISHL may be considered as long as all the other physical causes are ruled out. An emotional disorder may alter the neurovascular mechanism due to an ANS response. There would be vascular contractions, viscosity alterations and eventual congestion, anoxia, increase in capillary permeability and local metabolic impairment, what would justify a SHLThe intense anguish and restlessness during consultation and the great variability in results are factors that suggest a psychosomatic SHLNoteworthy and disproportional recovery after clinical treatment.The fact that the cochlea is mainly supplied by one single artery, the labyrinthine artery, and this is a terminal artery, makes the inner ear very much prone to circulatory alterationsa)Total and permanent vessel occlusion;b)Total and temporary vessel occlusion;c)Cochlear blood hypo flow.In vascular causes, the problems may be present on the blood vessel wall, as is the case in arteritis and spasms, or intravascular, such as gaseous emboli, fat emboli, polycythemia, hyperviscosity , sickle cell crisis, and others. Cochlear damage would be secondary to anoxia or hypoxia generated and could occur due to three different mechanismsTotal and permanent occlusion of a blood vessel would cause anoxia, and consequently necrosis of labyrinthine membranes, fibrosis and ossification of the inner ear. This would justify the hearing loss, but not the recovery of tonal thresholds. Moreover, rare are the times in which fibrous and bone proliferation were found in some study. Thus, this is not a very probable theory that could justify the ISHL pathogenesis.Cochlear tolerance to schemia is very limited and the action potential starts to be impaired after 60 seconds of anoxiaLow blood flow, the third mechanism involved, happens because of a hyperviscosity, generating a drop in oxygenation (insufficient to maintain cochlear metabolism) and cochlear hypo function. This would be the most acceptable theory among vascular causes, since it does not only justifies HL, but also the possible audiological recovery and even cochlear histological findingsBlood flow is inversely proportional to blood viscosity. Viscosity depends on the hematocrit, serum viscosity, red cell aggregation and RCD. RCD is defined as a physicochemical characteristic that allows the red blood cell to pass through capillaries with diameters that vary between 3 and 12µm, although in average their diameter is always above 7µm. It is believed that a reduction in RCD may cause some cochlear injuries and be related to SHLThe hematocrit plays a more important role in the blood flow of the great vessels, while RCD plays an important role in the circulation blood flow, specially in smaller vessels, and it may be affected under some situations such as coronary artery diseases, blood diseases and cellular shape changes, renal disease, post-operative alterations and post AMI alterations, serum protein alteration, peripheral vascular disease or diabetes mellitus. It is also thought that smoking may have influence over RCDIt is believed that RCD reduction may be either an acute or chronic phenomenon, and it bears great incidence in cases of ISHL. A schemia secondary to RCD reduction could be clinically mistaken with an obstructive vascular cause and explain the hearing loss, and moreover a possible recovery of tonal thresholds in some cases of ISHL. Hence the use of drugs that could enhance blood viscosity and RCD would be justifiable.Still, we know that an acute upper airway infection, together with a viremia, alters RCD and can lead to an ischemia on the vascular stria due to a low blood flow and increase in arterial-venous shunt; both the ischemia and acidosis generated worsen RCD reduction, thus causing a vicious cycle. Thus, if an upper airway acute infection reduces RCD and is associated to a viremia, the ISHL pathogenesis may associate vascular and viral factors as ISHL originAllergic factors, despite rare, may also cause changes in the cochlear microcirculation or, by anaphylactic phenomena, and trigger a change in capillary permeability. Therefore, the physiopathology in these cases is really more related to a vascular alteration then the allergic cause itselfThe fact of the matter is that SHL under blood hyperviscosity is highly controversial. Being that because of a reduction in blood flow in the capillaries that nurture the stria vascularis and consequently cause ischemia in the inner ear, or a reduction in the oxygen transporting capacity because of a slower blood flow and consequent generation of micro thrombi and lesion to the cochlear endothelium, thrombosis to the venous system of the inner ear; or any other hypothesis, it is clear that there is no absolute way to proveThe inner ear hemorrhage could also be an important explanation for ISHL. Secondary to the use of anticoagulants or diseases that cause low platelet levels, such as leukemia and aplastic anemia, traumas, meningitis, subarachnoid hemorrhage and adenocarcinoma metastasis, this bleeding would then cause a sudden pressure rise in the endolymph or perilymph, biochemical and osmolarity alterations in the intracochlear fluids, intracochlear conduction disorders and ischemia of the peripheral region next to the bleeding site. Histological alterations found in the cochlear of these thrombocytopenic patients show and prove such bleeding in the peri and endolymphatic spaces, besides cellular infiltrate. Notwithstanding, considering that the perilymphatic space hemorrhage is responsible for SHL, the mechanisms are not as clear as they seem, since many arguments contradict them. We did not notice any blood-induced hearing loss in the perilymph during stapedectomy; no histology study show damage to the cochlear structures caused by the presence of blood in the perilymphatic space after stapedectomy; and even after the experimental infusion of blood in the swine perilymph; no potential amplitude or threshold alteration was noticed on the electrocochleogram. Therefore, not even an insidious hemorrhage in the perilymphatic space could cause SHL: thus bringing about a huge stalemate. Despite a not very clear physiopathology, there seems to be an intimate relation between the aforementioned diseases and the hearing lossStill, among vascular causes, it is known that western diets offer a greater risk of ISHL when compared to the Japanese diet, since ours is more loaded with saturated fat. Western diet causes higher platelet aggregation and factor VII activity, increasing blood coagulability and the risks of vascular obstruction. Now, the eastern diet, richer in vegetal oils, has more important antithrombotic and antiatherosclerotic properties that prevent vascular diseasesThus, a history of vascular disease or platelet or red blood cell alterations are the only clinical findings that could lead us to infer a vascular cause for ISHL; Histopathologically, we should find fibrous and bony proliferation in the cochlear and semicircular canal spaces, or at least, the hemorrhage in the cochlear spaces, proven by not only image studies, but also by post mortemBecause of the presence of antibodies against antigens in the inner ear and the formation of immune complexes in the stria vascularis, endolymphatic sac and duct, an autoimmune disease may be consider a cause that favors sudden or progressive HLAutoimmune SHL may happen alone – specific organ-disease – as in the thyroiditis of Hashimoto, or associated to one systemic disease – non-organ-specific disease, such as Cogan's syndrome, nodose polyarthritis, systemic lupus eritematous, rheumatoid arthritis, Berget's disease, Sjögren's syndrome, Behcet's syndrome, ulcerative colitis, rheumatoid arthritis, and othersThe immune reaction in the inner ear caused by the endolymphatic via is more intense than that in the middle ear. It is characterized by cell infiltration  and, after three weeks, T suppressor cells coming from the blood circulation. There still is an inflammatory reaction with increase in local antibodies and intracochlear fluids, with consequent damage to the inner ear , besides an intimate involvement of the endolymphatic sac: if destroyed it reduces the immunogenic response. During the secondary response, the endothelial cells of the spiral modiolus vein are activated, thus taking on the characteristic of a differentiated endothelial vessel, and recruit lymphocytes of the blood circulationAlthough the labyrinth immunity be similar to that of the brain , the inner ear immunity seems to be more immunoresponsive, because the antigens present in the cochlear quickly stimulates the systemic immunitySometimes, immune responses may happen not directly related to an autoimmune disorder or antigen, but rather due to the use of a drug or antibody.Toubi et al. (1994) found a growing prevalence of anticardiolipine antigen (aCL) in patients with ISHL, without any association with autoimmune disorders. It is known that this antibody bears important association with vascular thrombosis and is not always correlated to clinical alterations; its presence may be secondary to some viral infections and it probably justifies the hearing loss, although we need more studies, based on larger samples, to confirm it. Thus, SHL cause would not be purely caused by an autoimmune alteration, but rather triggered by a viral infection or vascular disorder. This is another factor against autoimmune disorders causing ISHL.Apha-interferon (a-IFN), on its turn, is a drug used to treat acute and chronic viral infections, neoplasias and autoimmune disorders, and it generates side effects such as psychiatric disorders, blood alterations, autoimmune disorders , they submitted that the rupture of one of the labyrinthine windows  would cause the loss of perilymph and consequent pressure alteration in the relationship between compartments with perilymph and endolymph. In 1986, Kohut et al. and Kamerer et al. in 1987, apud Yoon et al. (1990), described the presence of a labyrinthine fistula causing micro fissures between the niche of the round window and the posterior semicircular canal ampule. With that, there would be a rupture of the Reissner membrane and the fluid mixture, with consequent cochlear hypo function, but not a broad cochlear dysfunctionMoreover, no rupture of the cochlear windows, or either Reissner or basilar membrane rupture was seen in any of the studies carried out by Schuknecht & Donovan (1986) and Yoon et al. (1990) that would justify such theory.It is important to bear in mind that such occurrence usually happens after sudden pressure alterations in the middle ear, as are the cases of head injuries, barotraumas, stapedectomies or intense physical exercises and not spontaneouslyThe viral infection, due to a number and considerable circumstantial evidences is currently considered as one of the major causes of ISHL. The broad association with the histological findings on temporal bone, seroconversions of specific viral titles, and in a lesser degree, but not less important, connection with prior history of a flue or another prior viral disease is proof of such fact.Considering the virus as a possible cause, this suggests that the agent responsible has a broad distribution in the human body, besides affinity for the nervous tissue. Either the acute or the latent attack may then generate damage to the inner ear and consequent hearing lossUpper airway infections (UAI) and intracranial infections may reach the inner ear through the blood (stria vascularis) and through the cochlear aqueduct or internal acoustic meatus IAMAlthough acute viral infections may cause such damage, latent infections and their reactivation may also explain the lesion. The main latent viruses are part of the group of herpes virus: they are ubiquitous, bear strong neurotropism as characteristic; they not always cause symptoms  and have complex relation with ISHL. They are herpes simplex, herpes simplexOther agents are known causes of hearing damage also in fetuses and newborn babies, such as the ones from syphilis, toxoplasmosis and rubella - STORCH (syphilis/ toxoplasmosis/ rubella/ CMV and herpes virus)Almost all the SHL cases caused by virus are unilateral. The bilateral occurrence is rare. Depending on the viral agent, the hearing loss may present a certain pattern. For example, mumps courses with severe and irreversible HL, while HL caused by VVZ is of low severity and reversibleDue to the cochlear arterial blood supply and venous drainage, there should be a correlation between cell degenerationsThe lack of correspondence between the structural degenerations  thus suggest that the ganglionic cells of the apical turn are the primary target of the causal factor. Therefore, an isolated lesion on the ganglionic cells of the apical turn without loss of corresponding sensorial structures is not compatible with the theory of vascular ischemia as a cause for SHL, and much less with membrane rupture, but it is rather compatible with the viral origin.Despite the fact that cochlear vascularization is equivalent in normal ears and ears with ISHL, it is noticed that there are more histopathological alterations on cochleas affected by ISHL than in normal cochleas, or even on those affected by presbicusisHowever, although we see many similarities as to anatomopathological alterations among the ears affected by ISHL and ears affected by viral infections, some evidences are still contradictory as to the fact that the virus may be a causal factor of this clinical manifestation.It is not always possible to establish a direct causal relation with the virus, because the antibody titles may not be significantly altered in a SHL supposedly caused by a viral agentMoreover, the levels of MxA, an a-IFN protein marker of systemic viral infection in leucocytes no always present significant increase in patients with ISHLEven then, the fact that the structural lesions found in temporal bones of ISHL patients be very characteristic and similar to the ones found in temporal bones affected by a viral infection contribute to this theory  as the main etiology for ISHLHaving said that, we notice that autoimmune diseases, and specially membrane rupture, seem not to have important correlation with idiopathic sudden hearing loss. Vascular disorders and viral agents, however, are the causing factors most likely to justify this hearing loss, although a prior vascular disease or viral exposure is not necessarily inherent to the origin of the HL. ISHL low incidence, the difficult access to the inner ear for better studies on unknown diagnosis and physiopathology make it difficult to achieve better conclusions as to the very origin of this clinical manifestation.


Inonotus hispidus mushroom is a traditional medicinal fungus with anti-cancer, antioxidation, and immunomodulatory activities, and it is used in folk medicine as a treatment for indigestion, cancer, diabetes, and gastric illnesses. Although I. hispidus is recognized as a rare edible medicinal macrofungi, its genomic sequence and biosynthesis potential of secondary metabolites have not been investigated. In this study, using Illumina NovaSeq combined with the PacBio platform, we sequenced and de novo assembled the whole genome of NPCB_001, a wild I. hispidus isolate from the Aksu area of Xinjiang Province, China. Comparative genomic and phylogenomic analyses reveal interspecific differences and evolutionary traits in the genus Inonotus. Bioinformatics analysis identified candidate genes associated with mating type, polysaccharide synthesis, carbohydrate-active enzymes, and secondary metabolite biosynthesis. Additionally, molecular networks of metabolites exhibit differences in chemical composition and content between fruiting bodies and mycelium, as well as association clusters of related compounds. The deciphering of the genome of I. hispidus will deepen the understanding of the biosynthesis of bioactive components, open the path for future biosynthesis research, and promote the application of Inonotus in the fields of drug research and functional food manufacturing. Inonotus hispidus (Bull.) P Karst. (Hymenochaetaceae) is a well-known edible and medicinal mushroom with a long history as a health food and ancient folk medicine in Europe -6,7-vinyl]-3,5-dioxafluoren-5-one (5), inoscavin C (6), hypholomine A (7), inoscavin E (8), inonoblin A (9), inonophenol A (10), inonophenol B (11), hispolon (12), hispinine (13), methyl 5--3-hydroxypenta-2,4-dienoate (14), MBP (15), interfungin C (16), interfungin A (17), inonophenol C (18), inonotusin A (19), inotolactone B (20), eburicoic acid (21), hispindic acid B (22), 3β-hydroxy-lanosta-8,24-dien-21-al (23), inonotusol F (24), inonotusol G (25), inonotusane F (26), cerevisterol (27), 4,6,8(14),22(23)-tetraen-3-one-ergostane (28), 7(8),22(23)-dien-3-one-ergostane (29), inonotsutriol E (30), inonotsutriol A (31), inonotusane E (32), inotolactone A (33), and inonotusol E (34) (1–19) and the lanosterol-type triterpenoids (20–34), which correspond to the distinct clusters formed in the network  , Table 3 network , Table 3 species .Sanghuang is a well-known macrofungal medical herb in China, Japan, Korea, and other Asian countries. It has a long history of medicinal usage and health benefits [Compendium of Materia Medica. Indeed, sanghuang is recorded in various medical documents as a generic term for a group of medicinal macrofungi with specific biological activity and similar morphology, which is difficult to reconcile with modern species classification systems, seriously limiting modern medicinal research on sanghuang. Although the establishment of Sanghuangporus genus containing 15 species [Sanghuangporus species, several medicinal fungi from the genera Inonotus and Phellinus, such as I. hispidus [P. gilvus [sanghuang in traditional medicine. Such fungi are more aptly known as sanghuang-like fungi. These valuable medicinal macrofungi should not be excluded from the Chinese medicinal fungus sanghuang because of the development of Sanghuangporus genus. The ability of genome sequencing to reliably correlate morphological traits to the genome enables the differentiation of sanghuang-like fungi from Sanghuangporus species, which is difficult to separate morphologically.benefits ,67,68, w species ,69,70 hahispidus  and P. g. gilvus , are stiI. hispidus have traditionally been used as medicines for the treatment of indigestion, cancer, diabetes, and gastric diseases by the residents of Xinjiang province and Northeast China [1) [2) [15) [19) [20) [I. hispidus are essentially polyphenols [I. hispidus. Indeed, polyphenolic compounds with a styrylpyrone backbone are abundant in the genera Phellinus and Inonotus [Sanghuangporus [Sanghuangporus species. Furthermore, the molecular network constructed based on GNPS clearly displays the differences in the chemical composition of the fruiting bodies and mycelium of I. hispidus. This may serve as a precise guide for the targeted isolation of specific compounds  , phellib [1) [2) , MBP (15[2) [15) , inonotu19) [20)  (Figure yphenols , which aInonotus , as wellangporus , reflectompounds . The molompounds .G. lucidum identified multiple biosynthetic genes necessary to produce ganoderic acids, which provided vital information to uncover the biosynthetic pathways of these essential medicinal components [A. cinnamomea yielded critical candidate gene information to unravel antrocamphins biosynthesis in phase II clinical trials. Although polyphenols with a styrylpyrone moiety are a characteristic component of sanghuang-like fungi, the biosynthesis for these chemicals has received little attention [I. hispidus indicated that BGC12, with g1457 as a core gene, and BGC13, with g1510 as a core gene (Genome sequencing of mponents . The genttention ,71,75. Core gene , Table 2I. hispidus is thought to be related to the multiple P450s spread throughout the chromosomes, notably the over 50 P450 members of unidentified gene families (Secondary metabolite biosynthesis genes, especially post-modification genes, found in mushrooms tend to be scattered across chromosomes, similar to plants, rather than clustered like bacteria. The presence of high oxidation of polyphenolic compounds and triterpenoids in families .I. obliquus, a homology of I. hispidus, has more than 20 sesquiterpene synthases [I. hispidus only has 15 sesquiterpene synthases and one sesquiterpenoid xylaritriol [I. hispidus are inactive.Basidiomycetes-derived sesquiterpenes are a class of important natural products with diverse structures and various activities, and sesquiterpene synthases are a class of critical genes for natural product biosynthesis . Accordiynthases  and eighynthases , but I. aritriol , suggestInonotus hispidus is a well-known medicinal mushroom that exhibits anti-cancer and immunomodulatory activities, as well as a long history of usage as a medicinal fungal material with various health benefits. Here, for the first time, we provide the de novo assembled complete genome of I. hispidus. Chromosome-level assembly and functional annotation described in this study provide useful clues for subsequent gene functional research. According to comparative genomic analysis, the genus Inonotus has different gene compositions. Phylogenomic and evolutionary analysis of the genus Inonotus reveals evolutionary traits. The investigation of mate locus and CAZyme facilitates artificial cultivation. A thorough examination of secondary metabolite biosynthesis genes showed a wide range of biosynthetic potential. Further molecular network-based metabolite analysis revealed differences in chemical composition and concentration in fruiting bodies and mycelia. This work not only covers a vacuum in I. hispidus genetic information but also gives crucial insights into the biological aspects of the medicinal-edible fungus I. hispidus, such as growth characteristics and biosynthesis routes of bioactive components. A thorough grasp of I. hispidus’ genome will pave the way for its future application in pharmacological research and functional food development. In short, the genome sequencing of I. hispidus sheds light on the biosynthesis and medical applications of its metabolite.