This study aimed to evaluate the effectiveness of and satisfaction with hearing aids as a treatment option for tinnitus with hearing loss. Methods: This retrospective study used the tinnitus handicap inventory (THI), the satisfaction with amplification in daily life (SADL) questionnaire, and a medical chart review. A total of 116 patients treated between August 2018 and December 2020 were included. All patients with tinnitus and hearing loss underwent the same counseling sessions. Sixty patients chose to have hearing aids fitted (aided group), whereas 56 patients chose not to (non-aided group). Both the groups had similar audiometric configurations, durations of tinnitus, and ages. Structured interviews were performed, with various measures evaluated using the visual analog scale (VAS) and the THI questionnaire, before and six months after fitting the hearing aids. The SADL questionnaire was administered 6 months after fitting the hearing aids. Results: The patients’ THI scores reduced 6 months after the counseling, but the improvement in the THI scores was only significant in the group that received hearing aids. There were significant differences between the VAS scores of the two groups, and the changes in the VAS scores in the groups were statistically different. Subjective satisfaction with a hearing aid increased with improvements to tinnitus-related discomfort. Conclusion: The study’s results indicated that patients with hearing loss and tinnitus can be treated with hearing aids and counseling. Tinnitus is the perception of sound by a person without the presence of an external sound source. Various reasons can cause tinnitus, and its treatment is challenging because it has a deep association with the central nervous system and emotion. Acoustic therapy is important for treating tinnitus. This therapeutic method enables patients to decrease their recognition of tinnitus by increasing external auditory stimuli using a hearing aid or a sound generator . Since sHearing aid technology has developed significantly over the past years, but limited studies have investigated improvements in tinnitus following hearing aid use in patients with hearing difficulties. According to Surr et al., 62% of patients who began using a hearing aid had tinnitus, which declined in approximately half of them following hearing aid use . This apTherefore, this study aimed to (1) compare improvements in the THI scores of patients who choose hearing aids with counseling and in those who choose only counseling and to (2) investigate the association between improvements in tinnitus and subjective patient satisfaction with hearing aids after the tinnitus improvements based on the THI score and the SADL questionnaire.Our data were obtained from patients with bothersome tinnitus and hearing loss treated at the tinnitus clinic in our medical center between August 2018 and December 2020; 60 patients were treated with hearing aids and counseling (aided group), and 56 patients were prescribed only counseling (non-aided group). No information about this study was provided to the patients. Patients were excluded if they received other treatments, used a combination of devices, experienced other potential confounding injuries  during the study, had a low THI score (<15) at their first visit, or did not use their hearing aids as recommended. Patients with vertigo were also excluded from our study, and patients with asymmetrical sensorineural hearing loss received an MRI to exclude other diseases, such as vestibular schwannoma. Tinnitus etiologies were distinguished from other middle-external ear diseases and Meniere’s disease through otolaryngological and neurological examinations. No patients had ever received counseling as a treatment, and none had previously used hearing aids. The study protocol was approved by the Institutional Review Board of Medical Center (IRB No. 2019-07-065).This study was a retrospective, questionnaire-based evaluation of treatment success. All first-visit patients underwent audiological evaluations. Pure-tone audiometry was performed at a frequency range of 0.25–8 kHz in a sound-proof room using an interacoustic AC40 audiometer. BC audiometry was performed at 0.5, 1, 2, and 4 kHz frequencies. Speech discrimination scores were calculated by estimating the percentage of correct word recognition out of a list of 50 phonetically balanced words, which was presented at 40 dB louder than the speech reception threshold. All the patients in the study were tested using audiological evaluations, REM, and hearing aid fittings performed by two clinicians with extensive experience. All the patients received counseling, and the test results were explained. As a treatment option, amplification was recommended for each patient. In the study, the counseling group chose not to use hearing aids or other sound therapy devices. Counseling was provided by an otologist with extensive experience in tinnitus management. Counseling was performed based on the Jastreboff tinnitus model and treatment strategy. The counseling can be summarized as follows: (1) an explanation of tinnitus’s origins, (2) a description of the tinnitus evaluation and hearing test, (3) a resolution of questions and anxiety about tinnitus, and (4) an explanation of the treatment course and goal of tinnitus management.Hearing aid selection depended on personal preference and audiometric configuration. All the hearing devices used in this study were digital hearing aids. For patients who had unilateral hearing loss with tinnitus, we recommended a unilateral hearing aid on their affected side. Two hearing aids were recommended if the patient had bilateral hearing loss. When patients with bilateral hearing loss could only use unilateral hearing aids due to personal preferences or economic conditions, a unilateral hearing aid was applied to the side where tinnitus sounds were louder. The audiometric profile typical of most people suggested tinnitus at normal low frequencies with mild to moderate high-frequency SNHL; thus, hearing aids with open-fitting slim tubes were frequently selected. Every hearing aid was optimized for the amplification of low-input sounds. Real ear probe microphone measurements  were performed for all hearing aids to support optimal match prescriptions, and the prescription method was NAL-NL2. During the fitting period, the devices were further tuned according to subjective reports. For tunings, follow-up visits were scheduled based on each patient’s requirements. The hearing aid group was required to use their devices maximally, mostly in quiet environments where the tinnitus was very clear, even when communication was not a high priority.The self-administered THI questionnaire was used to quantify the impacts of tinnitus on individuals. They were classified as emotional stress, functional disorders, and disaster thoughts. The questionnaire had 25 questions, and each had the following responses to choose from: “Not so”, “Sometimes it is so”, and “It is so”, which were scored as 0, 2, and 4 points, respectively. We measured THI scores before either the counseling or hearing aid fittings and 3 months after the first THI measurement.For the satisfaction investigation, we used the SADL scale, which consisted of 15 questions covering four items  6 months after hearing aid use commenced. The SADL scale, which consists of a global score , was used, ranging from ”not at all” (1 point) to “always so or very much so” (7 points). One of these values was selected for scoring. The SADL investigation was performed three months after the hearing aid fittings. We conducted a follow-up fitting every month. The total VAS score was obtained based on the following question: “How loud or strong was your tinnitus on average over the last month?” The score ranged from 0 to 10 .t-test was used to compare the demographic data and VAS scores for tinnitus in the groups. The chi-square test was used to evaluate the effects of hearing aids on tinnitus according to changes in THI scores. The total changes in and classified THI scores of the two groups and the changes in their THI and SADL scores according to the fitting sites and the locations of tinnitus were analyzed with the Mann–Whitney test. The changes in the THI scores and the overall SADL scores were assessed using Spearman’s correlation test. Statistical significance was set at p < 0.05.The data were analyzed using SPSS software . Student’s All the patients had sensorineural hearing loss. In most cases, the patients had mild symmetrical high-frequency gently sloping hearing loss. The two groups showed no significant differences across the frequencies tested between 250 Hz and 8000 Hz for both ears .The differences between the THI scores obtained before and after wearing a hearing aid and receiving counseling (aided group) or receiving only counseling (non-aided group) were categorized as follows: >30 points, very much; 10–30 points, moderate; 1–10 points, a little; and <1 point, not at all. In the aided group, 51 (85%) patients reported that the hearing aids were “helpful to some degree”, and 9 (15%) reported that they were “not helpful at all”; these numbers for the non-aided group were 41 (73%) and 15 (28%), respectively .p < 0.05; p < 0.05; Each group showed significant improvements in the VAS scores, and the changes in the VAS scores of both groups  were significant , whereas 19 wore bilateral hearing aids . p < 0.05; The changes in the THI and overall SADL scores showed a positive correlation and were statistically significant , HHI, and APHAB are well-known questionnaires that investigate the effects of hearing aids ,24,25. HWe had several limitations in this study. Patients were not randomly assigned in this study, which was conducted through a chart review as a retrospective study. As a result, there may have been a selection bias in the sample selection process. Rachel et al. recently published a randomized controlled trial study in this field ,28. MoreIt is expected that the prescription of hearing aids for patients with hearing impairments and tinnitus can provide audiological benefits and can positively impact tinnitus treatments. In addition, the THI can be used to evaluate the degree of an audiological improvement to predict and evaluate success following hearing aid use in such patients, and the SADL questionnaire is helpful for measuring satisfaction by considering various variables.


Hearing loss is a sensory deprivation that can affect the quality of life. Currently, only rehabilitation devices such as hearing aids and cochlear implants are used, without a definitive cure. However, in chronic hearing-deprived patients, in whom secondary auditory neural degeneration is expected, a relatively poor rehabilitation prognosis is projected. Stem cell therapy for cochlear neural structures would be an easier and feasible strategy compared with cochlear sensory cells. Considering the highly developed cochlear implantation technology, improving cochlear neural health has significant medical and social effects. Stem cell delivery to Rosenthal's canal in an acutely damaged mouse model has been performed and showed cell survival and the possibility of differentiation. The results of stem cell transplantation in chronic auditory neural hearing loss should be evaluated because neural stem cell replacement therapy for chronic (long-term) sensorineural hearing loss is a major target in clinics. In the present study, we established a mouse model that mimicked chronic auditory neural hearing loss (secondary degeneration of auditory neurons after loss of sensory input). Then, mouse embryonic stem cells (mESCs) were transplanted into the scala tympani and survival and distribution of transplanted cells were compared between the acute and chronic auditory neural hearing loss models induced by ouabain or kanamycin (KM), respectively. The mESC survival was similar to the acute model, and perilymphatic distribution of cell aggregates was more predominant in the chronic model. Lastly, the effects of mESC transplantation on neural signal transduction observed in the cochlear nucleus (CN) were compared and a statistical increase was observed in the chronic model compared with other models. These results indicated that after transplantation, mESCs survived in the cochlea and increased the neural signaling toward the central auditory pathway, even in the chronic (secondary) hearing loss mouse model. Hearing loss is a sensory deprivation that can affect the quality of life. Hearing loss has significant social implications, and the association of hearing loss with cognitive dysfunction has been recently introduced . TherapeSound transduction is a complicated process performed by sensory and neural structures. Therefore, sensory and neural structures are well organized to properly transduce sound. When bypassing the sensory part among these structures, for example, using hearing aids or cochlear implants to augment hearing, maintaining the health of neural hearing structures remains important . Therefoin vitro differentiation or organoids has been performed [Stem cells, by definition, are cells that can proliferate and differentiate . Pluripoerformed –8. Howeverformed . The diferformed –13. In cStem cell delivery to Rosenthal's canal has previously been attempted. Stem cell survival in both Rosenthal's canal and perilymphatic space and the possibility of differentiation have been proposed and performed in acutely damaged mouse models . HoweverIn the present study, a mouse model was established that mimicked chronic auditory neural hearing loss (secondary degeneration of auditory neurons after loss of sensory input). Then, mouse embryonic stem cells (mESCs) were transplanted into the scala tympani and survival and distribution of transplanted cells evaluated and compared to the acute auditory neural hearing loss model induced by ouabain. Lastly, the effects of mESC transplantation on neural signal transduction, observed in the cochlear nucleus (CN), were evaluated.n = 4), kanamycin (KM) only (n = 5), KM plus stem cell (SC) (n = 6), ouabain only (n = 3), and ouabain plus SC (n = 3). In the ototoxic treatment groups, ototoxic agents were delivered only to their left ear . Before the surgery to induce secondary auditory nerve degeneration after ototoxic damage and mESC transplantation, the mice were anesthetized with an intramuscular injection of a mixture solution (0.1 mL/20 g) prepared by diluting a 1 : 3 mixture of normal saline from 3 : 1 mixture of Zoletil  and Rompun . All experimental surgeries including mESC implantation were performed in the left ear only. Hearing was measured using the auditory brainstem response (ABR) and assessed before and after ototoxic agent injection and mESC transplantation as shown in Twenty-one male, 5-week-old, adult C57BL/c6 mice were used in this study. The animals were divided into five groups: no treatment control (v/v) heat-inactivated fetal bovine serum (FBS) , 0.1 mM β-mercaptoethanol , 0.1 mM GlutaMAX (Gibco), 0.1 mM ES qualified nonessential amino acid (NEAA) , 1% penicillin-streptomycin (PS) (ATCC), 1000 U/mL leukemia inhibitory factor , 0.033% CHIR99021 , and 0.125% PD035901 (Tocris Bioscience) at 37°C in a 5% CO2 humidified incubator.The ESCs tagged with green fluorescent protein (GFP), donated by Prof. Hosup Shim , were used in this study. The same culture method for maintenance followed the previously published method due to the same donated cells . Cells wμL of a 1 mM solution) or KM (5 μL of a 150 mg/kg solution) was placed into the round window to deliver the drug through the scala tympani via a microcannula tube connected to a Hamilton syringe. After the surgery was completed, the incision was sutured and the mice stabilized on a warm pad until normal movement. To evaluate the hearing changes, the ABR was measured before and 2 weeks after ouabain treatment. After KM treatment, the ABR was measured before treatment and at 2, 4, and 8 weeks after treatment.Two auditory neural hearing loss mouse models were created. First, an acute model was designed to mimic acute neural hearing loss with intact sensory structures. Second, a chronic model was created to resemble the chronic status of sensorineural hearing with secondary neural degeneration resulting from the lack of sensory input from the cochlea. Ouabain  and KM  were used to develop the acute and chronic mouse models, respectively. To induce auditory neurodegeneration in the mouse, the posterior wall of the left ear was dissected until the bulla appeared. Next, the bulla was carefully removed until the round window of the cochlea was exposed. A Gelfoam soaked with ouabain (5 μL of mESCs (2 × 104 cells/μL) was injected into the scala tympani of the cochlea at 8 weeks after KM treatment. After transplantation, the incision was sutured in the same manner as in the method for inducing auditory nerve degeneration.For the mESC transplantation experiment, at 2 weeks and 8 weeks after auditory nerve degeneration induction in the acute and chronic mouse models, respectively, green fluorescent protein- (GFP-) tagged mESCs were transplanted into both models. The surgical technique followed a previously published report in a way that our research team continued it . In brieTo measure hearing changes, the ABR was performed with tone bursts and a specific stimulated frequency . The treated mice were placed in a sound chamber, and three needle electrodes were inserted at the vertex (active) and under both auricles (reference and ground). To evaluate the hearing threshold, tone burst stimulations of 8, 16, and 32 kHz were delivered through a tube inserted into the left ear of the mouse. The ABR waveform was measured by averaging 512 signals for 10 ms/step measured from 80 dB SPL to 10 dB at 5 or 10 dB intervals.μm thick sections and stained with hematoxylin and eosin (H&E). Then, the sections were observed under a microscope  to evaluate the histological changes in peripheral auditory organs after mESC transplantation in the auditory neural hearing loss model.Mice were subjected to cardiac perfusion under anesthesia to obtain the cochlea and brain. The cochlea and brain were harvested from mice in each group. The cochleae were postfixed in 4% PFA and then decalcified in 0.1 M EDTA solution for 4 days at room temperature (RT). The cochleae were washed with PBS for 3 h to stop the decalcification and embedded in a paraffin block. The blocks were sliced into 5 w/v) serial sucrose solutions until the tissue relaxed. Then, the brains were embedded in the OCT compound  and sliced into 5 μm thick sections under cold conditions using a microtome . The prepared slides were stained for epifluorescence imaging.The harvested brains were postfixed in 4% PFA overnight at 4°C, and crystallization was prevented by soaking in 10, 20, and 30%   and 0.3% Triton X-100 (Sigma-Aldrich) to prevent nonspecific binding. To identify GFP-positive cells inside the cochlea, the slides were incubated with neurofilament antibody  overnight at 4°C without GFP. The following day, the slides were washed three times with PBS for 5 min and then incubated with secondary antibody  for 1 h. Nuclei were stained using 4′,6-diamidino-2-phenylindole (DAPI).To evaluate vesicular glutamate transporter 1 (VGLUT1) expression in CN after mESC implantation in the auditory neural hearing loss model, samples were incubated with VGLUT1 antibody  overnight at 4°C. The next day, samples were washed as described above. Animals without any treatment were sacrificed as controls. Then, the samples were incubated with secondary antibody  for 1 h and visualized using DAPI. A confocal microscope  was used to obtain representative images. In the acquired images, the intensity of VGLUT1 within the CN was measured using ImageJ software 1.43u .t-tests. To analyze the VGLUT1 expression, the Kruskal-Wallis test, a nonparametric one-way ANOVA, was performed and Dunn's test for multiple comparisons was used as the posttest. A p value < 0.05 was considered statistically significant.The results were expressed as means ± standard deviation (SD) and analyzed using GraphPad Prism software  or SPSS software . The Kolmogorov-Smirnov test was used to determine whether data were parametric. Two-way ANOVA was performed as well as Bonferroni tests as posttests to analyze the ABR thresholds. Significant differences between the ototoxin-treated group and ESC-transplanted groups were statistically analyzed using two-tailed paired After ouabain (acute model) and KM (chronic model) injection, the ABR threshold in the ipsilateral ear increased significantly at all frequencies  .t-test: 8 kHz: t = 3.464, df = 2, p = 0.0742; 16 kHz: t = 1.000, df = 2, p = 0.4226; and 32 kHz: t = 1.000, df = 2, p = 0.4226)  (two-tailed paired  0.4226) .t = 13.34, p < 0.001; 16 kHz: t = 12.29, p < 0.001; and 32 kHz: t = 11.26, p < 0.001), indicating the possibility of chronic hearing deprivation. At 2 weeks after GFP-mESC transplantation into the cochlea through the round window in the chronic auditory neural hearing loss mouse model (8 weeks after KM treatment), ABR was measured to evaluate hearing improvement. However, compared with the chronic auditory neural hearing loss model, improvement in hearing threshold was not observed   rates of 8 kHz and 16 kHz were 78.6% and 92.8% for 32 kHz, respectively. The increased threshold remained unchanged until 8 weeks  . These rIn the acute or chronic auditory neural hearing loss model, the histological changes in the peripheral auditory organs were observed using H&E staining as shown in However, after mESC transplantation, cells (probably supporting cells or perhaps transplanted mESC) in the organ of Corti were observed in both acute (ouabain) and chronic (KM) neural hearing loss models . In RoseTo track the delivery pattern and survival of mESCs in the cochlea easily, mESCs tagged with GFP were transplanted into mice with both auditory neural hearing loss models. The GFP expression in peripheral auditory organs was observed in half of the mice in the chronic model and in all mice in the acute model at 2 weeks after mESC transplantation into the scala tympani through the round window. In both neural hearing loss models, mESCs were observed in all turns of cochlea. mESCs were observed in Rosenthal's canal in both neural hearing loss models. Obvious GFP expression was observed in mice in the acute model and weak GFP expression in mice in the chronic model. Limited GFP and neurofilament coexpression was observed, indicating relatively low differentiation of mESCs into neural structures. In mice in the chronic model, especially in the perilymph, transplanted mESCs formed cellular aggregates showing GFP-positive oval structures with varying densities. The structures extended into the scala vestibuli, which is anatomically distant from the initial transplantation site. In mice in the acute mouse model, several mESCs migrated into the organ of Corti and resided adjacent to supporting cells and tunnel of Corti . Based op < 0.001; ouabain vs. ouabain plus mESCs, p < 0.01; The connection between SGN and CN, which is the proximal central neural structure, was evaluated, and alteration of this connection due to mESC transplantation in both auditory neural hearing loss models was confirmed. The VGLUT1 expression in the dorsal cochlear nucleus (DCN) of the CN within the brainstem, the first structure to the central auditory pathway, was observed . The VGLExpecting a positive effect on secondary auditory nerve degeneration in the case of chronic sensorineural hearing impairment with a relatively long prevalence is difficult using cochlear implantation. Therefore, in the present study, mESCs were transplanted directly into the scala tympani, which is mainly composed of neural structures in Rosenthal's canal, as a method to repair or restore damaged nerves. Subsequently, whether the transplanted cells in the cochlea can survive or improve hearing function was investigated. Our established auditory neural hearing loss mouse models, induced acutely by ouabain and chronically by KM, showed increased ABR thresholds and decreased SGN density compared with controls. However, difference was observed in the morphology of the organ of Corti in peripheral auditory organs between the two models. Conversely, in the central auditory pathway, mice in the two models showed the same pattern, a decreased VGLUT1 expression in the CN. Improvement in hearing after mESC transplantation was not observed; however, the survival of transplanted cells in the cochlea was confirmed based on GFP-expressing cells in the cochlea and increased VGLUT1 expression in the CN. The mESC-transplanted mice in both models showed the cells probably supporting cells or perhaps transplanted mESC in the organ of Corti without flat epithelium and no decrease in SGN density. In particular, the distribution and morphology of GFP expression in cochlea were different in the two models. Although GFP-tagged mESCs were observed in all turns of cochlea and Rosenthal's canal in both neural hearing loss models, KM-treated mice in the chronic model formed cellular aggregates and showed weak GFP expression compared with ouabain-treated mice in the acute model. This result indicated that the transplanted mESCs can survive in the perilymphatic region and Rosenthal's canal and their potential to differentiate in a chronic auditory sensorineural hearing loss model.In the present study, a neuropathy model was made by delivering ototoxic reagents (ouabain and KM) into the cochlea via the round window. In a prior report, ouabain-induced neuropathy showed normal levels of DPOAEs and EP, indicating the normal function of outer hair cells and stria vascularis, and selectively induced SGN loss . AlthougTwo methods can be suggested for the mechanism of repairing damaged cells or tissues through stem cell therapy. One is that stem cells directly engraft and differentiate into required cell types. The other is that the transplanted donor cells produce a paracrine effect, which stimulates the recipient cells to restore the damaged tissue on their own. Studies of the paracrine effect show that mammalian tissues have reproducible cells and only need to accept appropriate signals to initiate regeneration and repair. In addition, donor-transplanted cells have a long-term effect on tissue regeneration, even though they are short lived. In other words, the presence of the transplanted stem cells is important to initiate tissue repair, but once the patient's own cells are activated, the transplanted stem cells are no longer needed.A previous study has been reported using umbilical cord blood cells to treat spinal cord injury. The results of this study showed that the transplanted stem cells only existed for 7 to 10 days, but compared to injured animals that did not receive cord blood cells, wound lesions were reduced and the mobility of spinal cord-injured mice was significantly improved .Reportedly, in the acute model, transplanted mESCs are possibly differentiating to the neural lineage without any additional treatment, even outside Rosenthal's canal. The target lineage includes glial cells and astrocytes which are considered supporting cells for maintenance of neural structures in the brain . The difThe present study results indicate the possibility of clinical application of stem cell therapy for hearing rehabilitation. However, regeneration or recovery of damaged neural structures in mouse cochleae was not observed. Considering the short duration of mESC transplantation (2 weeks) and use of pluripotent stem cells, which should be differentiated in multiple steps, the study period was insufficient to observe the transdifferentiation of stem cells to compensate for complex sound transduction systems. Nevertheless, the delivery of stem cells with low efficiency and increase of central signal transduction, possibly through the paracrine effect of pluripotent cells, could be a meaningful result. To substantiate these results further, additional studies are necessary. First, the trophic factors responsible for the paracrine effect should be identified. Second, molecular characteristics of the transplanted cells in different parts of the cochlea should be further analyzed. The additional experiments and confirmation of safe stem cell delivery  could support regeneration of neural structures to improve hearing rehabilitation in the near future.The survival of transplanted cells in the cochlea was confirmed based on GFP-expressing cells in the cochlea and increased VGLUT1 expression in the CN. In particular, the distribution and morphology of GFP expression in the cochlea was different in the two models. Although GFP-tagged mESCs were observed in all turns of cochlea and Rosenthal's canal in both neural hearing loss models, KM-treated mice in the chronic model formed cellular aggregates and showed weak GFP expression nearby neural structures compared with ouabain-treated mice in the acute model. Improvement in hearing after mESC transplantation was not observed; however, the survival of transplanted cells in the cochlea was confirmed based on GFP-expressing cells in the cochlea and increased VGLUT1 expression in the CN. This result raises the possibility of using stem cell replacement therapy for hearing rehabilitation in patients with chronic nerve damage.


Azaphilones are a family of polyketide‐based fungal natural products that exhibit interesting and useful bioactivities. This minireview explores the literature on various characterised azaphilone biosynthetic pathways, which allows for a proposed consensus scheme for the production of the core azaphilone structure, as well as identifying early diversification steps during azaphilone biosynthesis. A consensus understanding of the core enzymatic steps towards a particular family of fungal natural products can aid in genome‐mining experiments. Genome mining for novel fungal natural products is a powerful technique for both exploring chemical space and providing new insights into fungal natural product pathways. Azaphilones are a polyketide‐based family of fungal natural products with a highly oxygenated bicyclic pyrone‐quinone structure, many of which have useful and interesting bioactivities. This minireview reveals the common steps in azaphilone biosynthetic pathways to aid the use of genome mining to identify orphan azaphilone biosynthetic gene clusters. They can react readily with amines to form vinylogous γ‐pyridones, hence the name ‘azaphilone’ Figure , A.1,1,2 Az1,2;3.This minireview focusses on the biosynthesis of azaphilone compounds and compares common biosynthetic steps between this family of natural products. We examine the biosynthetic pathways to several related compounds (where their biosynthesis informs studies on azaphilones): the important mycotoxin, citrinin C‐methyltransferase) domain  Figure S1).6, 7 T.6, 7 22.11 is a well‐known mycotoxin that is closely related to the azaphilones, but citrinin 1 itself is resistant to amination.1 biosynthesis used radioisotopes, which confirmed its polyketide origin through the incorporation of 14C‐labelled acetate.[1 biosynthesis through a series of gene knockout and heterologous expression experiments  responsible for the biosynthesis of asperfuranone 2, a compound which is closely related to the azaphilones, consisting of a bicyclic furan containing core , homologous to citS, and a highly reducing PKS . Deletion of either PKS abolishes asperfuranone 2 biosynthesis, leading to the proposal that the hrPKS produces a reduced starter unit 9 that is transferred to the SAT domain of the nrPKS to be further extended.2 BGC identified from Aspergillus terreus  with homology to salicylate monooxygenases. In the biosynthesis of asperfuranone, the FMO AteAfoD hydroxylates C‐4 of 8, leading to spontaneous cyclisation and dehydration to give the shunt azaphilone 10  and a hrPKS (azaB)  demonstrated that is solely responsible for the flavin‐dependent conversion of the benzaldehyde intermediate FK17‐P2a 19 into the bicyclic, pyran‐containing azaphilone compound, azanigerone E 17 , in contrast to the asperfuranone pathway, led to the accumulation of azanigerone E 17. This suggested that AzaB is only responsible for the biosynthesis of the 2,4‐dimethylhexanoyl chain 20, which is proposed to be the substrate for the O‐acyltransferase AzaD to esterify the C‐4 hydroxyl group of azanigerone E 17 to produce azanigerone B 14. Additional predictions were made about the pathway, based on a time course experiment and isolation of the metabolites, but not further verified.The first BGC encoding azaphilone biosynthesis was reported by Tang and co‐workers in 2012.) Figure , A, whicer fungi.14, 152.4Chaetomium globosum, for example chaetoviridin A 2425.C. globosum by searching for clustered homologues of the nrPKSs and hrPKSs present in the asperfuranone and azanigerone BGCs  or the hrPKS (cazF) abolished production of any azaphilone compounds, with no other intermediates accumulating. Knock‐out of the O‐acyltransferase, cazE  accumulated cazisochromene 26 . Cazisochromene 26 is likely to be the product of C‐4 hydroxylation by CazL and C‐6 chlorination by CazI of cazaldehyde A 27. Timing of chlorination versus hydroxylation is unclear. Interestingly, no serine hydrolase is present in this BGC, suggesting that its activity is not always essential in azaphilone biosynthesis. In vitro assays using CazF (hrPKS), CazE (O‐acyltransferase), and cazisochromene 26, resulted in the production of chaetoviridin A 24. This is in accord with the hrPKS giving the 5‐hydroxy‐4‐methyl‐3‐oxohexanoyltriketide 28 that is required to produce the lactone ring fused to the isochromenone core of chaetoviridin A 24. The lack of any azaphilone compounds produced by the ΔcazF strain suggested that the hrPKS is also implicated in the production of the 4‐methyl‐hex‐2‐enoyltriketide starter unit 29 required by the nrPKS (CazM).in vitro assays of CazF and CazM produced cazaldehyde A 27, the first enzyme free intermediate that undergoes hydroxylation and chlorination before cyclisation and dehydration forms the azaphilone core. Experiments showed that the SAT domain only recognised the more reduced triketide.Characterisation of a second azaphilone pathway was reported in 2012 by Tang and co‐workers,2.5Monascus spp. such as monascorubrin 30 and rubropunctatin 31 was reported in 2013 by Kwon and co‐workers , which is homologous to other known azaphilone nrPKSs, was targeted for gene deletion in M. purpureus, to create strain ΔMpPKS5. This strain no longer produced any azaphilones, demonstrating that MpPKS5 is essential to azaphilone biosynthesis in M. purpureus. Kwon and co‐workers19 in a similar manner to citrinin 1 and asperfuranone 2 biosynthesis.[Monascus ruber studied by Chen and co‐workersSaccharomyces cerevisiae did not increase yield compared to expression of MrPigA alone.32 to give 19.[3 cluster, although the same ketoreduction does occur, and Tang and co‐workers speculated that AzaE is instead responsible.19 to form azanigerone E 17  abolished T22 azaphilone 38 biosynthesis, with the deletion strain accumulating harziphilone 39 and its isomers 40 and 41. The authors proposed that the biosynthesis proceeds in a similar manner to that of the chaetoviridins and chaetomugilins  being produced as a shunt compound during asperfuranone 2 biosynthesis  require an nrPKS to biosynthesise a benzaldehyde intermediate, the titre of which may be increased by the serine hydrolase . The nrPKS may require an hrPKS to produce a specific reduced starter unit for further extension.The current consensus Scheme  for the s Figure , B, furts Figure , B, as wThe FAD‐dependent monooxygenase catalyses C‐4 hydroxylation, which is the key step in azaphilone biosynthesis, converting the benzaldehyde into the bicyclic, pyran‐containing core of azaphilones. It is interesting to note that two mechanisms have been suggested for the biosynthesis of this core structure. Wang and co‐workersO‐acyltransferase. The acyl moiety can be biosynthesised by either an hrPKS  (Table A clinkerT) Table .O‐acylation step is not necessary for the formation of the bicyclic pyrone‐quinone azaphilone core), or alternatively using the MpFas2/MrPigJ‐K fatty acid synthase dimer sequences instead of AzaB, or using diversifying enzymes sequences, for example CazI, the halogenase from the chaetoviridins and chaetomugilins BGC.To test this theory, we have conducted a representative cblaster4O‐acyltransferase) see Table Fungal genome mining is an exciting method to help realise the full natural product potential revealed within fungal genomes. Many tools exist to facilitate fungal genome screening, two of the most popular are fungiSMASHThe authors declare no conflict of interest.Dr. Katherine Williams received her PhD from the University of Bristol (UoB) in 2010, before moving to the Bristol Polyketide Group, investigating the biosynthesis of fungal metabolites with interesting bioactive properties. Research posts at the Leibniz Universität Hannover, Germany with Prof. Russell Cox, and at Cardiff University with Prof. Ruedi Allemann followed. She now works in the UoB, with Dr. Andy Bailey and Prof. Chris Willis, on a project aimed at developing a high‐throughput heterologous production platform for fungal natural product antibiotic discovery.Dr. Claudio Greco received his PhD at the University of Bristol (2017) elucidating the biosynthesis of fungal secondary metabolites under the supervision of Prof. Russell Cox and Prof. Chris Willis. This was followed by a two‐year postdoctoral research at the University of Wisconsin‐Madison working with Prof. Nancy Keller, studying secondary metabolism regulation in pathogenic fungi. He is currently working at the John Innes Centre with Prof. Barrie Wilkinson as a BBSRC Discovery Fellow investigating the ecological role of fungal natural products.Dr. Andy Bailey is a senior lecturer at the University of Bristol, UK. His research interests are based around different aspects of fungal biology and their analysis using molecular genetic approaches. This includes genome mining to explore fungal secondary metabolism, fungi as pathogens of plants and invertebrates and other fungi, plus establishing methods for genetic analysis of basidiomycetes.Prof. Chris Willis is Professor of Organic Chemistry and Head of Organic and Biological Chemistry at the University of Bristol. Her research focuses on natural product biosynthesis including the application of total synthesis, isotopic labelling, pathway engineering and mechanistic studies to produce biocatalysts and new bioactive molecules. She was the recipient of the Natural Product Chemistry Award of the Royal Society of Chemistry in 2020.As a service to our authors and readers, this journal provides supporting information supplied by the authors. Such materials are peer reviewed and may be re‐organized for online delivery, but are not copy‐edited or typeset. Technical support issues arising from supporting information (other than missing files) should be addressed to the authors.Supporting InformationClick here for additional data file.


Rheumatoid arthritis (RA) is a chronic inflammatory disease which is marked by leukocytes infiltration inside synovial tissue, joints and also inside synovial fluid which causes progressive destruction of joint cartilage. There are numerous genetical and lifestyle factors, responsible for rheumatoid arthritis. One such factor can be cysteine cathepsins, which act as proteolytic enzymes. These proteolytic enzyme gets activated at acidic pH and are found in lysosomes and are also termed as cysteine proteases. These proteases belong to papain family and have their elucidated role in musculoskeletal disorders. Numerous cathepsins have their targeted role in rheumatoid arthritis. These proteases are secreted through various cell types which includes matrix metalloproteases and papain like cysteine proteases. These proteases can potentially lead to bone and cartilage destruction which causes an immune response in case of inflammatory arthritis. The histological hallmarks of rheumatoid arthritis gets characterised via the development of aggressive pannus which invades inside bones and cartilage. The joint parenchymal lesions are detected with synovial hyperplasia and cellular infiltrate which contain numerous cells like monocytes/macrophages, lymphocytes as well as neutrophils. The abundantly present cells inside synovial fluid and tissues are macrophages/monocytes, exerting crucial role in pathogenesis of RA . It lead2Rheumatoid arthritis is an auto-immune disorder. The complete mechanism by which it occurs is not fully elucidated but it is observed that in patients with RA, autoantibodies are produced against the components of extracellular matrix (ECM). This in turn causes destruction of immune system which in combination with synovial cells leading to disease progression. Diarthro-dial synovial joint comprises of particular connective tissues as well as a fibrous capsule. 70% of the material of bone is of inorganic nature , organic material denotes 20% (comprising of type I collage), and 10% contains water . The bonCathepsins can be defined as the heterogeneous group of proteases which are found inside lysosomes and possess acidic environment. Cathepsin is derived from a greek work Kathepsein which means to digest. Cathepsin family comprises of 15 members, which are divided on the basis of their catalytic activity. The three major groups of their distinction include serine proteases (contains cathepsin A and G), aspartate proteases . Most of the cathepsin (expect E and G), reside inside endosomal compartment. Hence, they are termed as lysosomal cathepsins. These are termed as intracellular active enzymes which are accountable for proteolysis inside acidic environment, where they degrade intracellular as well as extracellular proteins. They also exert some functions under normal conditions which are shown in The activity of cysteine cathepsin is based on their pH as well as their cellular localization. The characterization of cysteine cathepsin reveals that it comprises of cysteine residue at the active site. Their structure is homologous to that of papain (which is a cysteine protease obtained from the papaya fruit) and thus are termed as papain like cysteine protease. Cathepsin which are in combination with parent protease papain are termed as MEROPS which acts as peptidase database. They are expressed inside vertebrates, primitive parasites, plants, and invertebrates. They get transferred inside lysosomes through a specific pathway (mannose-6-phosphate pathway) and can play a potential role in chronic as well as infectious disease. Matured proteolytic active cathepsin gets released after their activation via the removal of N-terminal propeptide, which occurs at low pH of lysosomes . The var3Cathepsin G also termed as CTSG belongs to the family of serine proteases, which was initially detected inside azurophillic granules of neutrophil leucocytes. They are also detected inside myeloid cells, including primary human monocytic cells, myeloid dendritic cells, B cells, murine microglia and plasmacytoid dendritic cells. CTSG is a 255-amino acid residue protein, on undergoing cleavage it leads to formation of 2 amino acids and 11 amino acids residue at N- and C-terminal side of pro-CTSG. Initially it gets stored inside the primary granules and after being stimulated via immune complexes or through pharmacological agents CTSG gets released to the extracellular space. Cathepsin G exert its numerous functions as it can remove pathogens and can regulate inflammation via attenuating the level of cytokines, chemokines and cell surface receptors. They have become a new target for biomarkers and thus they can serve as the target for auto-immune disease . They leCathepsin G can alter the cell shapes which brings about an intercellular gap between endothelial and epithelial cells, thereby enhancing their permeability. Possible mechanism of action involves attenuating calcium balance which causes amplified inositol phosphate level and thus activates protein kinase C leading to amplified albumin flux across cell membrane, ultimately causing cleavage of endothelial cadherin (which maintains vascular integrity). Evidences have shown neutrophil surface bound proteases causes cleavage of vascular endothelial cells via recruitment of neutrophils. It causes enhanced permeability of type II epithelial monolayers, further leading to structural changes and intercellular gaps which can be observed through scanning electron microscopy . Paracel4Initially, cathepsin K was extracted as cDNA using osteoclast library of rabbit, which is termed as OC-2. Later on three different group were isolated for human homologues which were termed as cathepsin K, X and O2. Predominantly, cathepsin K is present in abundant amount in skeleton (mainly inside osteoclasts). Systematic analysis has shown the observed role and action of cathepsin K inside human skeleton. They have their central action inside osteoclastic resorption of bone matrix. Cathepsin K is abundantly present inside osteoclast. To lesser extent, it is observed inside human heart, skeletal muscle, ovary, testes, small intestine, lung, and colon. Cathepsin inhibitors like leupeptin was able to inhibit the process of resorption inside the osteoclast cell assays. The cells treated with cysteine protease inhibitor comprises of vacuoles which contained collagen fibrils in undigested form. This indicates that degradation of collagen is initiated via cysteine protease . Cathepsperi-vascular interstitial matrix. Amplified expression of cathepsin K is associated with Larsen score which reflects the destruction and radiological changes inside joints. The level of cathepsin K-derived type I collagen telopeptides don’t have a specific correlation with bone degradation and its loss. The above data supports a potential role of cathepsin K in destruction of articular cartilage as well as subchondral bone. The various factors which induce production of cathepsin K inside the synovium comprises of TNF-α and IL-1. Cathepsin K respond towards IL-1 as well as TNF-α and enhances their production. Enhanced production of cathepsin K can lead to the invasion of RA synovial fibroblast at pannus cartilage. Thus, cathepsin K serves its valuable action for the evaluation of joint destruction in patients suffering from rheumatoid arthritis and the potential candidate which can inhibit cathepsin can serve as potential therapy, thereby preventing joint destruction  causes enhanced resorptive activity and thus contributing towards bone as well as cartilage erosion. Expression and activity of cathepsin K around lymphocytic infiltrate causes facilitated transportation of mononuclear cells via truction .5Cathepsin S serve as potent protease comprising of wide range of substrate including elastin, albumin, and insulin B-chain and hemoglobulin. Out of all these, high elastolytic activity of cathepsin S serve its major role in rheumatoid arthritis. Enzyme elastase serve its potential activity inside extracellular matrix at neutral pH. Cathepsin S is found in dendritic cells, B cells and in macrophages. The higher expression of cathepsin S is observed inside lungs where it can serve its role in lung diseases. Lower transcriptional levels of cathepsin S are ascribed to have their role in osteoarthritis multinucleated giant cells which are responsible for removing synovial bone debris. The above finding may indicate the role of cathepsin S in extracellular matrix degradation. Cathepsin S exert its potent proteoglycan degrading action, thereby serving its role in hydrolysing the aggrecans at acidic as well as neutral pH. It is expressed inside synovial macrophages in patients with rheumatoid arthritis. The secretion of cathepsin S inside cartilage matrix during rheumatoid arthritis can serve as an inflammatory process which can be deleterious. This results in alteration of integrity of the aggrecan type II cartilage and collagen. Cathepsin S is a critical protease required in antigen presentation. On comparison with Cathepsin K, it is observed that cathepsin S acts as weak collagenase and thus the degradation/damage via cathepsin S is lesser as in relation to cathepsin K .Cathepsin L belongs to cysteine protease and is abundantly expressed inside thymic cortical cells and are assumed to serve significant role for potential selection of T cells which must recognise non-self-structures. The expression of cathepsin L is triggered via proto-oncogenes (c-Harus and c-fos). It is expressed inside synovial cells where it attaches with bone and cartilage, leading to erosion. Cathepsin L leads to the degradation of proteoglycans as well as collagen compartments (including collagen type IX and XI), thereby promoting bone resorption and contributing towards cartilage and bone destruction. They also interfere with framework system of inflammatory cytokines, mediating the proteoglycan degradation via interleukins (mainly interleukin-1). It facilitates the migration of blood-borne mononuclear cells into synovium, which becomes the most pathogenic factor in conditions like rheumatoid arthritis. Cathepsin L also degrades type IV collagen which acts as major component of basement membrane and also degrades the cartilage components. Cathepsin L exert its major effect in rheumatoid as when compared to osteoarthritis, which is based on the fact that it binds and attaches to hyperplastic synovial lining to the bone. This phenomenon is only observed during rheumatoid arthritis. The degradation of matrix can occur via invasion inside pannus tissue. Thus, from the above outcomes it can be concluded that cathepsin L exert its role in rheumatoid arthritis .Another cathepsin which can serve its role in rheumatoid arthritis includes Cathepsin D. Cathepsin D is abundantly produced via chondrocytes and exert its enhanced activity in rheumatoid arthritis. Cathepsin D acts via degrading the central core protein of proteoglycans. This degradation takes place at acidic pH. Literature reports have also suggested that cathepsin D can process pro-active cathepsin L and B into their targeted active forms. Amplified action of cathepsin L and D is observed when both acts simultaneously as this potentiate/augments action of cathepsin L (i.e. enhanced transvascular migration of cells inside the synovium). Cathepsin D serve its potential role via the process of invasion as well its correlation with level of c-myc. From numerous studies it can be demonstrated that altered level of matrix metalloproteinases (such as stromelysin and collagenase) could serve its potential role in rheumatoid arthritis .6From numerous studies it can be reported that pharmaceutical inhibition of cysteine cathepsin can prevent the progression of bone and cartilage destruction in rheumatoid arthritis. The basis for treatment of rheumatoid arthritis includes non-steroidal anti-inflammatory drugs as well as corticosteroids. Proper understanding and advancement in the region of cytokine network can be serve as the base for identification of targeted therapies. Although, targeted therapies can’t cure rheumatoid arthritis but can help in the prevention of the disease progression. The action of cathepsin can be inhibited via implication of cysteine protease inhibitor which are designed as site-directed low molecular weight compounds. The active site of cathepsin proteases comprises of a nucleophillic thiol residue which can be counteracted by using an electrophillic moiety . This warhead can be placed into a peptide/missile section which gets recognised by substrate binding region of protease. Interaction of missile takes within S-binding sites . The type of reaction (reversible or irreversible bonding) depends on the type and nature of electrophillic moiety. Irreversible inhibitors that have high potency as well as selectivity are not considered to be suitable drug candidates for treatment of chronic disease like arthritis and osteoporosis. The fact of not using them is that their reactivity will be continuous i.e. it will react over and over again with numerous reactive cysteine protein species leading to toxic/lethal side effects. This can lead to generation of immunogenic haptens which are produced as a result of formation of covalently bound inhibitor-cathepsin adducts .Under physiological conditions, action of cathepsins is regulated via the presence of endogenous inhibitors which are termed as cystatins. The most widely implicated cystatin is cyastatin C which acts extracellularly. It serves as cysteine inhibitor which exert its wider action and is linked with arthritis. It can act via treating the chronic and severe arthritic cartilage lesions which can in turn decrease its progression. Down-regulation of cystatin C can lead to articular bone and cartilage damage. Over-expression as well as enhanced levels of cystatin C have their role in rheumatoid arthritis . Cathepshttp://www.novartis.com). The compound is given via oral route and dosage given is 10–50 mg/day. The reports of phase II demonstrated that it inhibits breakdown of collagen and thus improves bone formation (http://www.gsk.com). Numerous cathepsin K inhibitors have been synthesized and explored on the basis of their stability studies as well. Along with this their capacity to inhibit bone resorption was also studied. Peptide aldehyde inhibitor, inhibits cathepsin K and further it inhibits osteoclast mediated bone resorption inside human assays and animal assays. This improves bone and cartilage loss as well. The major problem with synthesis of cathepsin K inhibitors are mainly because of the lack of tissue specificity. The drug carrier such be such that it could deliver cathepsin inhibitor to the specific site . The inhibition can also be brought about by inhibiting the lysosomal compartment which can inhibit cathepsin K secretion as well the inhibition of type II collagen occurs via lysosomal compartment. Inhibiting lysosomal compartment inhibits both these processes. The other targeted therapy and strategy can be via implication of polymer conjugates. Polymer conjugates are low molecular weight inhibitors and acts as water soluble polymer carriers. Polymer conjugates are of greater importance and advantage because of their accumulation inside bone and cartilage targeted moieties such as in hard tissues like arthritic joints. The clinical studies have evaluated the efficacy of two polymer bounded inhibitors (mPEG-I as well as ST-PHPMA-I) which inhibits the activity of cathepsin K inside the synovial fibroblasts , Aventis, Novartis, Bayer, and Merck. Novartis under the clinical trials reported for the production/development of cathepsin K inhibitor (named as AAE581), which was implied for the treatment of rheumatoid arthritis and osteoporosis , which formulated an inhibitor of cathepsin S (MV57471). This served as a potential inhibitor and served its role in treatment of disease like rheumatoid arthritis, organ/graft rejection, multiple sclerosis, and pain. They can also serve their potential role in diseases like lupus and diabetes. Medivir lead to the development of another cathepsin S inhibitor which acts as a reversible inhibitor and serve its role in immune response towards antigens. Aventis and celera genomics are working on the development on cathepsin S inhibitors which can be the lead for treating numerous inflammatory and auto-immune disorders . The dru7Cathepsin are involved in diseases like rheumatoid arthritis, and inflammatory processes via matrix degradation. They are expressed inside immune system  which are involved in cartilage as well as bone turnover. Cathepsin K, S and G serve as major target for progression of rheumatoid arthritis via degrading collagen as well as extracellular matrix. The inhibition of cathepsin can lead to prevention of progression of diseases. Preclinical as well as clinical data encourages and supports the data that cathepsin inhibitors can serve as novel disease modifying and mechanism-based drugs. Action of cathepsins is regulated via the presence of endogenous inhibitors which are termed as cystatins. The most widely implicated cystatin is cystatin C which acts extracellularly. It serves as cysteine inhibitor which exert its wider action and is linked with arthritis. It can act via treating the chronic and severe arthritic cartilage lesions which can in turn decrease its progression.8This review received no funding from any agency of public, profitable or commercial sector.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.


Neuroinflammation, which is mediated by microglia and astrocytes, is associated with the progression of neurodegenerative diseases. Increasing evidence shows that activated microglia induce the expression and secretion of various lysosomal cathepsins, particularly during the early stage of neuroinflammation. This trigger signaling cascade that aggravate neurodegeneration. To date, most research on neuroinflammation has focused on the role of cysteine cathepsins, the largest cathepsin family. Cysteine cathepsins are primarily responsible for protein degradation in lysosomes; however, they also play a role in regulating a number of other important physiological and pathological processes. This review focuses on the functional roles of cysteine cathepsins in the central nervous system during neuroinflammation, with an emphasis on their roles in the polarization of microglia and neuroinflammation signaling, which in turn causes neuronal death and thus neurodegeneration. Neuroinflammation is a common mechanism that plays a crucial role in the pathogenesis of various neurodegenerative diseases ,2. PrimaGlia cells, consisting of microglia, astrocytes, and oligodendrocyte lineage cells, are superior to neurons in cellular diversity and function and are considered neuron-supporting cells . In the Peptidases have been identified as important players in chronic innate neuroinflammation mediated by microglia and astrocytes ultimately associated with the progressive nature of neurodegenerative disorders ,21. PeptLysosomal peptidases represent lysosomal hydrolytic enzymes that catalyze the breakdown of proteins ,31. TheyThe name cathepsin derives from the Greek verb katahepsein (“to digest”) and was first proposed for peptidases active in slightly acidic environments but was later used for all aspartic, serine, and cysteine cathepsins ,34. BecaThe most extensive group of peptidases are cysteine peptidases, which includes 11 cathepsins  . They beThe expression of cysteine cathepsins depends on tissue distribution and their biological function. Most cathepsins are ubiquitously expressed in human tissues, while others have more specific distributions. For example, cathepsin K is mainly expressed in osteoclasts, osteoblasts, and epithelial cells and is involved in bone resorption . CathepsCathepsins are synthesized as inactive preproenzymes. Cleavage of the signaling protein and N-glycosylation of the remaining pro-cathepsin occurs during the passage to the endoplasmic reticulum. Glycosylated pro-cathepsin then enters lysosomes via the mannose-6-phosphate receptor pathway, where it is converted to the active form by the acidic conditions, autocatalytically, or by other peptidases . OptimalAlthough all cathepsins are targeted to endosomes/lysosomes, they are not equally distributed among different tissues ,47. Varicathepsin B is the most studied cathepsin expressed in the CNS. It is found in different brain regions, preferentially in neocortical and hippocampal neurons [Of all the cathepsins,  neurons ,53. Its  neurons  and cell neurons . It enga neurons ,48. Incr neurons  and the  neurons . Catheps neurons  and -11  neurons  and clea neurons . It play neurons ,62,63 an neurons . In chro neurons ,66,67.Cathepsin L is abundantly distributed throughout different brain areas and is preferentially expressed in neurons [ neurons , astrocy neurons . In the  neurons . It clea neurons . It is a neurons . Extrace neurons  and is a neurons .cathepsin H in pro-neuropeptide processing has been demonstrated. Cathepsin H has been associated with neuropeptide-producing secretory vesicles due to its cleavage of N-terminal basic residues of (Met)enkephalin [A novel role for kephalin . Converskephalin . In humakephalin . It is dkephalin . Apart fkephalin , a recenkephalin .Cathepsin C is expressed in a variety of mammalian tissues, with the highest levels in lungs, kidneys, liver, and spleen, but relatively low levels in the brain. The distribution of cathepsin C in normal brain is restricted to the neurons of the limbic system and several nuclei of the brainstem. Its granular immunohistochemical signals were found in neuronal perikarya of particular brain regions, such as the accessory olfactory bulb, septum, CA2 of the hippocampus, part of the cerebral cortex, medial geniculate, and inferior colliculus [lliculus ,78. Cathlliculus .Cathepsin S is unique within the cysteine cathepsin family due to its ability to retain activity at a neutral pH, which increases its potential involvement in extracellular proteolytic processes [rocesses ,81. It irocesses . Cathepsrocesses ,48,82,83rocesses ,85. For rocesses . Cathepsrocesses . Furtherrocesses , includirocesses .cathepsin X. Cathepsin X is an important cysteine peptidase in degenerative processes during normal aging and in neurodegenerative diseases. It is primarily expressed in cells of the immune system, such as monocytes, macrophages, and dendritic cells, but also widely expressed in brain cells, which implies its involvement in neuroinflammatory processes. In the brain, it is localized in microglia, astrocytes, aged neurons, and even oligodendrocytes [Another cathepsin that has been studied for its role in inflammation-associated neurodegenerative diseases is drocytes ,46,89. Wdrocytes ,90. Addicathepsin K expression was also observed in multiple neurons as well as in astrocytes and white matter oligodendrocytes [Apart from all of the mentioned roles of cathepsins in CNS, drocytes . Severaldrocytes ,92. It idrocytes . Furtherdrocytes . Cathepsdrocytes .Inflammatory processes in the CNS are directly involved in neuronal death and therefore in the development of neurodegenerative diseases such as Alzheimer’s and Parkinson’s disease. Microglia are the resident innate immune cells in the brain and actively contribute to neuronal damage in neurodegenerative diseases . Various1-40 and Aß1-42 [1-40 and Aβ1-42 are two of several forms of Aβ that have the same N-terminal but differ in their C-terminal residues [One of the major proteins responsible for the formation of amyloid plaques and toxic fragments in AD is neurotoxic Aβ, a polypeptide formed by the proteolytic cleavage of amyloid precursor protein (APP)  in secred Aß1-42 ,101. Aβ1residues .1-42 [In addition to the pathogenic role of cathepsin B in aging and age-related neurodegeneration, cathepsin B seems to degrade Aβ via C-terminal truncation, thus leaving its involvement in Aβ metabolism unclear ,103. It 1-42 . This em1-42 .Besides cathepsin B, also cathepsins L and S have been identified to cleave the wild-type β-secretase site . CathepsThe localization of cathepsins is not restricted to only intracellular compartments but can also be found in the extracellular space, indicating their broad spectrum of biological activities ,108,109.Upon stimulation with inflammatory mediators, cathepsin S can be released from activated microglia ,48,86. ACathepsins also contribute to damaging neurons and oligodendrocytes by their proteolytic action on axons and myelin proteins ,111. It Neuronal death is normal during nervous system development but can become devastating if not regulated, e.g., neuronal degeneration in chronic neurodegenerative diseases. It can be divided into necrosis and apoptosis and is regulated by several proteins and signal-transduction pathways. Players in the cell death and cell survival orchestra include: Fas receptor; Bcl-2 and Bax cytochrome c; caspases; p53; and extracellular signal-regulated protein kinases . LysosomAs already discussed, cathepsins are either localized intracellularly (within lysosomes) or extracellularly  and are c that causes apoptotic caspase activation [c exits mitochondria through these pores and, once in the cytosol, activates caspase-9 and thus promotes the executioner caspases-3 and -7 [Cathepsins are also involved in caspase activation and thus neuronal degradation by cleaving Bid protein. The cysteine peptidases B, H, K, L, and S are involved in cleaving pro-apoptotic Bid protein, a member of the Bcl-2 family, leading to the mitochondrial release of cytochrome tivation ,116. Bid3 and -7 . The maiCathepsin H is involved in lysosomal protein degradation ,119. Thelike cells induced by 6-hydroxydopamine by activating the caspase cascade [like cells [Additionally, cathepsin X promotes the apoptosis of neuron- cascade . Substanke cells . OverallAccumulating evidence suggests that neuroinflammation mediated by microglia and astrocytes is involved in the progressive nature of neurodegenerative disorders . NeuroinOveractivation of microglia and excessive amounts of released pro-inflammatory cytokines by microglia might result in neurotoxic consequences in neurodegenerative diseases. Increasing evidence indicates that microglial activation is an early and ongoing event in several neurodegenerative diseases . SeveralSimilarly, the expression of microglia-secreted cathepsin C is enhanced during CNS inflammation. Cathepsin C expression in the brain is predominantly induced in activated microglia , and micAltered cathepsin S expression is controlled by a built-in molecular clock in cortical microglia, and the circadian expression of cathepsin S is involved in diurnal variations of neuronal synaptic strength via proteolytic modification. Cathepsin S has also been associated with some sleeping disorders, as its genetic ablation causes reduced synaptic strength during sleep by inducing hyperlocomotor activity, which is required to obtain novel information after waking .substantia nigra pars compacta (SNc) of patients with Parkinson’s disease [Cathepsin L is also reported to play important roles in neuroinflammation-induced neurodegeneration. Cathepsin L is widely distributed throughout the CNS and is involved in the activation of microglia, an important source of cathepsin L . Catheps disease . Substan disease .Cathepsin H importantly contributes to peripheral inflammatory pathologies . It has Another cysteine cathepsin with an inflammatory role in the CNS is cathepsin X . The expDuring inflammation, M1-polarized microglia, evoked by exposure to IFN-γ or bacterial toxins , are characterized by an ameboid shape, high mobility, strong phagocytic activity, the production of pro-inflammatory mediators , and an increased expression of surface markers , which fuel the inflammatory process . In vitrRecent studies have shown that cathepsin X is also strongly associated with microglia polarization towards the neurotoxic phenotype. The proteolytic activity of cathepsin X in culture supernatants of activated microglial cells can be evoked by LPS stimulation. Upregulated expression and increased release and activity of microglial cathepsin X can lead to neurotoxicity mediated by microglia activation ,42. We dAlternatively, microglia can assume an M2 phenotype evoked by IL-4 or IL-13 that is characterized by thin cell bodies and branched processes. This could improve phagocytotic function and release numerous protective and trophic factors, triggering anti-inflammatory and immunosuppressive responses . IL-4-stA growing body of evidence shows that TLRs, which recognize a wide variety of danger signals and activate inflammatory cascades  and theiPro-inflammatory cytokines, TLRs, and other stress-like stimuli activate NF-κB , a trans2+-dependent protein kinase C/p38 MAPK/NF-κB cascade controls a range of cellular processes, including chemotaxis, phagocytosis, and cytokine secretion [Cysteine cathepsins have also been linked to the mitogen-activated protein kinase (MAPK) signaling pathway in microglia. The inflammatory response elicited by activated microglia is associated with MAPK activation, and this can lead to a variety of physiological processes, such as cell growth, differentiation, and apoptotic cell death . The MAPecretion . Additioecretion . Furthermore, p38 MAPK is activated in damaged areas in many neuroinflammation-related diseases, including lysosomal storage diseases. In Niemann-Pick disease type C (NPC), which is characterized by intracellular accumulation and redistribution of cholesterol in several tissues, including the brain , cathepsIt is widely accepted that neuroinflammation is an important factor in the pathogenesis of several neurodegenerative diseases and that the process is driven by activated microglia, which release pro-inflammatory mediators into the neuronal environment. Microglia-derived cysteine cathepsins are recognized as important pro-inflammatory mediators, triggering signaling pathways in inflammation-related cascades. Distinct cathepsins are upregulated in brain cells in the CNS during neurodegenerative pathologies associated with inflammation. Certain cysteine cathepsins have been found to be highly expressed and secreted from activated microglia, where a more detailed role of cathepsins in the microglia polarization towards M1 phenotype has been proposed. As cysteine cathepsins exhibit increased expression, activity, and subsequent secretion from activated microglia and participate in neuroinflammation-induced neurodegeneration, these peptidases are identified targets for the development of new diagnostic and therapeutic interventions in patients with neurodegenerative diseases. The beneficial effects of cystatins, endogenous cathepsin peptidase inhibitors, have yet to be demonstrated in neurodegenerative pathologies. Nevertheless, as they are general inhibitors, i.e., not cathepsin-specific, they can be expected to show off-target side effects. Therefore, rather than cystatins and other endogenous cysteine peptidase inhibitors, studies to date have focused on small synthetic inhibitors of appropriate specificities, with an emphasis on inhibiting excessive proteolytic activity of cysteine cathepsins associated with neuroinflammation-induced neurodegeneration. The design of selective and reversible cathepsin inhibitors is expected to improve peptidase-targeted therapy, which could significantly improve the treatment of patients with neurodegenerative disorders.


Certain consonant sounds called fricatives  are difficult to hear over the telephone; phones exclude high-frequency sounds that affect their intelligibility. This may be problematic for older adults responding to phone-based memory tests. Many older adults have some degree of hearing loss, and older men have it more in the high-frequency range. Hearing loss, in combination with phone bandwidth restrictions, may reduce older adults’ recall of fricative words. Participants  in the 1998 wave of the Health and Retirement study (HRS) completed a word list immediate recall task over the phone. List 4 recall was examined because it was evenly split (5 each) between words with and without fricative consonant sounds. Subjective ratings of hearing and health, age, depression, and education were also measured. A Wilcoxon signed-rank test showed participants recalled fewer fricative (M=2.8) than nonfricative (M=3.0) words, Z=-8.47, p<.001. An ordinal regression for fricative word recall indicated a sex by hearing interaction; males with worse hearing were less likely to recall more fricative words, OR=.94, 95% CI , p=.076, after controlling for age, education, health, and depression. An ordinal regression for nonfricative word recall did not show a main effect for hearing or a hearing by sex interaction. For both models, age, education, and health were related to recall. Consonant sounds may influence phone-based word recall, particularly for older men. Attention should be paid to word selection when designing phone-based cognitive tests in order to avoid memory impairment overestimation.


Evaluating the current health state in chronic otitis media (COM), audiologic results are complemented by subjective outcomes, such as health-related quality of life (HRQoL). Two disease-specific instruments assessing HRQoL in COM in German-speaking patients exist, i.e., the chronic otitis media outcome test (COMOT-15) and the Zurich chronic middle ear inventory (ZCMEI-21). Since the psychometric properties of these questionnaires in a concurrent application are unknown, the aim of this study was to compare the COMOT-15 and the ZCMEI-21.HRQoL was assessed in adult COM patients using the COMOT-15 and the ZCMEI-21. Psychometric properties were determined, including response distribution, concurrent validity, internal consistency, correlation to hearing and gender differences.r = 0.36, p < 0.0001; ZCMEI-21: r = 0.34, p < 0.0001).In 173 patients (mean age 51.5 years), both questionnaires showed normally distributed scores without strong floor and ceiling effects. The total scores and subscores of both questionnaires exhibited satisfactory internal consistency (Cronbach’s α 0.7–0.9) with the exception of the COMOT-15 hearing subscore (α = 0.94) and the ZCMEI-21 medical resource subscore (α = 0.66). Fair correlations between the air conduction pure-tone average and the total scores were found (COMOT-15: In the first study comparing the COMOT-15 and the ZCMEI-21, both questionnaires exhibited satisfactory psychometric properties with several subtle differences. The COMOT-15 has a strong focus on hearing with a probably redundant content of the hearing subscore and may be suited for hearing-focused research questions. The ZCMEI-21 provides a comprehensive assessment of the COM symptom complex and may therefore also be used in research settings, where ear discharge, vertigo or pain should be covered. The most frequent causes of conductive hearing loss are chronic inflammatory diseases of the middle ear, such as chronic otitis media (COM) with persistent discharge due to a chronic tympanic membrane perforation with or without cholesteatoma. Without adequate treatment, COM symptoms such as hearing loss or ear discharge can severely impair health-related quality of life (HRQoL) –3. When Validated and standardized questionnaires are used to assess the psychosocial impairment caused by hearing disabilities and accompanying symptoms such as tinnitus , 22. RegAdult patients with COM before surgical intervention were recruited from two tertiary hospitals . The study protocols were approved by the local Ethics Committees in accordance with the Helsinki declaration . Informed consent was obtained from all the participants.0.5–3 kHz) according to the Committee on Hearing and Equilibrium guidelines [0.5–3 kHz of the bone conduction (BC) threshold and the AC threshold.All audiometric measurements were performed with calibrated instruments in a sound-proof room (DIN EN ISO 8253). Measurements included standard pure-tone audiometry, performed with a clinical audiometer  in 5 dB steps. Pure-tone average (PTA) of the air conduction (AC) was calculated from AC thresholds at 0.5, 1, 2, and 3 kHz  or 95% confidence interval (95% CI) or as absolute numbers with percentages.All statistical tests were selected before data collection. Statistical analyses were performed using Microsoft Excel  and Prism . The significance level was set to γ and kurtosis κ were assessed. A normal (Gaussian) distribution is characterized by a skewness of 0 and a kurtosis of 0.Response distribution was assessed by determining the floor and ceiling effect, i.e. the percentage of patients exhibiting the lowest or highest possible score. Further, the skewness U test.Internal consistency as an indicator of reliability was assessed using Cronbach’s α . An α ≥ A total of 173 patients with a mean age of 51.5 years completed both the COMOT-15 and the ZCMEI-21. Detailed demographics and patient characteristics are given in Table The COMOT-15 and the ZCMEI-21 generally showed normally distributed scores  and the ZCMEI-21 total score .The COMOT-15 and the ZCMEI-21 strongly correlated in the total score . Although similar trends were observed in the ZCMEI-21, no statistically significant differences between men and women were found , the respective subscore was expected to strongly correlate with the AC PTA. Further, it was anticipated that it would correlate more strongly with the AC PTA than the respective ZCMEI-21 subscore, which assesses the psychosocial impairment by the entire symptom complex of COM, rather than solely focusing on hearing impairment. In accordance to these assumptions, a trend towards a higher correlation of the COMOT-15 psychosocial subscore and the AC PTA than the respective correlation of the ZCMEI-21 psychosocial subscore was found. This correlation may be weakened by the notion that a majority of patients perceive hearing impairment as the preponderant symptom among the symptom complex of COM. Consequently, hearing predominantly impairs HRQoL. In line, a slightly higher correlation between the COMOT-15 total score and the AC PTA was found compared to the respective correlation of the ZCMEI-21 total score and the AC PTA. As a clinical consequence, the COMOT-15 may be suitable for research settings that focus on hearing impairment or hearing improvement, such as ossicular chain reconstruction or implantable active hearing devices. In contrast, the ZCMEI-21 provides a comprehensive assessment of the COM symptom complex and therefore may also be used in research settings, where ear discharge, vertigo or (postoperative) pain should be covered .n = 3) of this subscore since Cronbach’s α is substantially influenced by the number of items [The ZCMEI-21 medical resource subscore showed a high floor effect indicating rather few demands of medical resources in the investigated cohort. In contrast, the COMOT-15 medical resources subscore (consisting of one question) showed more ceiling effects. This effect may be explained by the ZCMEI-21 medical resource subscore covering not only the number of consultations of an ENT-specialist, but also the use of local and systemic antibiotics. The COMOT-15 medical resource subscore only consists of a question assessing the number of consultations of an ENT-specialist and may therefore underestimate the use of medical resources of an individual patient. The Cronbach’s α in the ZCMEI-21 medical resource subscore was slightly below a satisfactory level of 0.7, which may be explained by the small number of questions  is not the only factor affecting HRQoL , 43. NotThis is the first study assessing information on the psychometric properties of the COMOT-15 and the ZCMEI-21 in a concurrent application. This study showed that both questionnaires exhibit satisfactory psychometric properties with several subtle differences. The COMOT-15 has a strong focus on hearing. However, its hearing subscore was found to be probably redundant with high floor and ceiling effects. Moreover, the COMOT-15 hearing subscore revealed large gender differences which may finally lead to gender differences in the total score. Consequently, the COMOT-15 may be suited for hearing-focused research questions in COM. When applying the COMOT-15, a gender sensitivity needs to be anticipated. The ZCMEI-21 provides a comprehensive assessment of the COM symptom complex and therefore may also be used in research settings, where ear discharge, vertigo or (postoperative) pain should be covered . Yet, si


MAFLD was also identified as the most important classifier for the presence of colorectal adenoma in decision-tree and random forest analyses . Among the three subtypes of MAFLD, non-obese MAFLD was the sole independent factor associated with the presence of colorectal adenoma . Non-obese MAFLD was also the most important classifier for the presence of colorectal adenoma in decision-tree and random forest analyses . MAFLD, particularly non-obese MAFLD, is the most important factor associated with the presence of colorectal adenoma rather than NAFLD. Colonoscopy examination should be considered in patients with MAFLD, especially those who are non-obese.Colorectal adenoma is linked to metabolic dysfunction. Metabolic dysfunction-associated fatty liver disease (MAFLD) has a precise definition and three subtypes, including non-obese MAFLD. We aimed to investigate the impact of MAFLD on the prevalence of colorectal adenoma by comparing it to non-alcoholic fatty liver disease (NAFLD) in health check-up examinees. This is a multicenter retrospective study. We enrolled 124 consecutive health check-up examinees who underwent colonoscopy. NAFLD and MAFLD were present in 58 and 63 examinees, respectively. Colorectal adenoma was diagnosed by biopsy. The impact of the MAFLD definition on the prevalence of colorectal adenoma was investigated by logistic regression, decision-tree, and random forest analyses. In logistic regression analysis, MAFLD was identified as the only independent factor associated with the presence of colorectal adenoma (OR 3.191; 95% CI 1.494–7.070;  The prevalence of colorectal cancer is increasing, with over 1.8 million new colorectal cancer cases being reported each year . A multiAging, gender, and family history increase the risk of colorectal cancer; however, these factors are unchangeable . Many liRecently, a panel of experts from 22 countries proposed a new definition for the diagnosis of metabolic dysfunction-associated fatty liver disease (MAFLD) . The diaThe aim of this study is to investigate the impact of MAFLD on colorectal adenoma by comparing it to NAFLD in health check-up examinees.In all subjects, the median age was 59 years and the male ratio was 80.6% . The medn = 72), respectively. Patients overlapping NAFLD and MAFLD comprised 69.4% (50/72) of the subjects with fatty liver  3.191; 95% confidence intervals (CI) 1.494–7.070; In a decision-tree algorithm, MAFLD was identified as the most important classifier for the presence of colorectal adenoma. Colorectal adenoma was observed in 51% of subjects with MAFLD, while colorectal adenoma was observed in 25% of subjects in the absence of MAFLD B. The seIn a random forest analysis for the presence of colorectal adenoma, MAFLD was identified as the distinguishing factor with the highest variable importance. NAFLD, age, and sex were the second, third, and fourth distinguishing factors, respectively C.Patients’ characteristics for subtypes of MALFD are summarized in p ≤ 0.001; We analyzed 7 factors, namely, obese-MAFLD, non-obese MAFLD, T2DM-MAFLD, age, sex, alcoholic intake, and smoking in the stepwise procedure. In the stepwise procedure, non-obese MAFLD was only selected as an explanatory variable for the logistic regression analysis. Non-obese MAFLD was identified as the only independent factor associated with colorectal adenoma . The prevalence of NAFLD has been reported to be between 40 and 52% in subjects who undergo colonoscopy ,14,15. MMAFLD was an independent factor for colorectal adenoma in this study. Decision-tree and random forest analyses also revealed that MAFLD, but not NAFLD, was the most important factor for the presence of colorectal adenoma. Recently, several studies have shown the superiority of MAFLD over NAFLD for the identification of patients with significant hepatic fibrosis, cardiovascular event, and chronic kidney disease ,12,19,20MAFLD consists of three subtypes, namely, obese-MAFLD, non-obese MAFLD, and T2DM-MAFLD ,22. We fThere are several limitations to this study. First, this study is retrospective, with a small sample size. Second, all participants were of Asian ancestry. Third, we did not evaluate factors associated with colorectal cancer, including dietary habits and physical activity. Fourth, since all subjects were health check-up examinees, the prevalence of severe steatosis and advanced fibrosis was low in this cohort. Therefore, we could not evaluate the impact of severity of hepatic steatosis and fibrosis on the prevalence of colorectal adenomas. Fifth, we also could not evaluate the impact of MAFLD on colorectal cancer, because of the small number of patients with colorectal cancer in this cohort (2/126). Accordingly, further studies should be designed in an international prospective study with a large sample size to more comprehensively evaluate the effects of MAFLD on the development of colorectal adenomas/adenocarcinomas using various factors including lifestyle habits and hepatic steatosis/fibrosis using FibroMax indexes ,34,35.In conclusion, we found that MAFLD was the only independent factor associated with the presence of colorectal adenoma. In both decision-tree and random forest analyses, these results were confirmed, while NAFLD was not an independent factor. In addition, we identified that non-obese MAFLD was associated with the presence of colorectal adenoma among the three subtypes of MAFLD. Since the identification of colorectal adenoma is an effective strategy to reduce the death rate from colorectal cancer, we suggest that colonoscopy examination is better considered in patients with MAFLD, particularly those who are non-obese.This study was designed as a multicenter cross-sectional retrospective study in Japan. The protocol conformed to the ethical guidelines of the 1975 Declaration of Helsinki as reflected by prior approval from the institutional review board of Kurume University School of Medicine (ID 20114). This research was performed in accordance with relevant guidelines and regulations. An opt-out approach was used to obtain informed consent from patients, and personal information was protected during data collection.n = 130): (1) subjects who were 20 years old or greater, and (2) subjects who underwent colonoscopy examination from 2018 to 2020. Of these, we have excluded subjects with (1) incomplete colonoscopy examination (n = 1), (2) positive results of hepatitis B virus surface antigen (n = 1) or hepatitis C virus antibody (n = 2), and (3) colorectal cancer (n = 2). Thus, a total of 124 subjects were analyzed in this study (n = 2) or with an incomplete examination, defined as an endoscope not reaching the cecum as documented by a picture of the ileocecal valve (n = 1). Finally, 124 health check-up examinees were analyzed in this study.We enrolled health check-up examinees who met the following inclusion criteria at the Kurume University Hospital, Kumamoto Central Hospital, and Kurate Hospital in Japan from April 2018 to March 2020  2 in this Asian cohort and T2DM was defined as HbA1c ≥6.5% or specific drug treatment. Metabolic dysregulation was defined as the presence of at least two metabolic risk abnormalities: (1) Central obesity (waist circumstance ≥90/80 cm in men and women), (2) blood pressure ≥130 mmHg or specific drug treatment, (3) plasma triglycerides ≥150 mg/dL or specific drug treatment, (4) plasma HDL-cholesterol <40 mg/dL for men and <50 mg/dL for women or specific drug treatment, and (5) prediabetes (fasting glucose levels 100 to 125 mg/dL or HbA1c 5.7–6.4%) [MAFLD was diagnosed according to the criteria proposed in 2020 by an international expert panel . The criColonoscopy examination  was performed by endoscopists with experience of performing more than 1000 procedures. Subjects were given polyethylene glycol for bowel preparation according to the manufacturer’s instructions. A complete examination was defined as an endoscope reaching the cecum as documented by a picture of the ileocecal valve. All identified colorectal polyps were removed and were tubular adenomas, which were diagnosed by pathological findings. This is a retrospective study and diagnosis of colorectal adenoma had already been made at the entry of this study.In all colonoscopy examinations, the quality of bowel preparation was graded as good . The withdrawal time of the colonoscopy procedure was at least 6 min to minimize the chance of missing lesions . IncomplContinuous variables are expressed as median and range or number. Categorical variables are expressed as frequencies and percentages. The differences between groups were analyzed using the Wilcoxon rank-sum test for continuous variables and the Fisher’s exact test for categorical variables. A logistic regression model was used to identify independent factors associated with the presence of colorectal adenoma. Data are expressed as OR and 95% CI.p < 0.05 was considered to indicate statistical significance. Data were analyzed using the JMP Pro15 .A decision-tree algorithm was constructed to reveal profiles associated with the presence of colorectal adenoma as previously described ,40. A raMAFLD, particularly non-obese MAFLD, is the most important factor associated with the presence of colorectal adenoma rather than NAFLD. Colonoscopy examination should be considered in patients with MAFLD, especially those who are non-obese.