Pulmonary delivery of mucus-traversing PF127-modified silk fibroin nanoparticles loading with quercetin for lung cancer therapy

The mucosal barrier remains a major barrier in the pulmonary drug delivery system,as mucociliary clearance in the airway accelerates the removal of inhaled nanoparticles(NPs).Herein,we designed and developed the inhalable Pluronic F127-modified silk fibroin NPs loading with quercetin(marked as QR-SF(PF127)NPs),aiming to solve the airway mucus barrier and improve the cancer therapeutic effect of QR.The PF127 coating on the SF NPs could attenuate the interaction between NPs and mucin proteins,thus facilitating the diffusion of SF(PF127)NPs in the mucus layer.The QR-SF(PF127)NPs had particle sizes of approximately 200 nm with negatively charged surfaces and showed constant drug release properties.Fluorescence recovery after photobleaching(FRAP)assay and transepithelial transport test showed that QR-SF(PF127)NPs exhibited superior mucus-penetrating ability in artificial mucus and monolayer Calu-3 cell model.Notably,a large amount of QR-SF(PF127)NPs distributed uniformly in the mice airway section,indicating the good retention of NPs in the respiratory tract.Themicemelanoma lungmetastasismodel was established,and the therapeutic effect of QR-SF(PF127)NPs was significantly improved in vivo.PF127-modified SF NPs may be a promising strategy to attenuate the interaction with mucin proteins and enhancemucus penetration efficiency in the pulmonary drug delivery system.

Developments of mucus penetrating nanoparticles

Mucus can effectively protect the exposed mucosal surfaces due to its adhesive and viscoelastic properties.Most foreign particulates are efficiently trapped in mucus layers via steric obstruction and adhesion.Trapped particles are typically removed from the mucosal tissue within seconds to a few hours depending on their location sites.This article focuses on describing the tenacious mucus barrier properties,the strategies to investigate the interaction of nanoparticles with the mucus as well as the novel developments of mucus penetrating nanoparticles.

Mucosal bacterial microflora and mucus layer thickness in adolescents with inflammatory bowel disease

AIM:To assess the mucosa-associated bacterial microflora and mucus layer in adolescents with inflammatory bowel disease(IBD) .METHODS:Sixty-one adolescents(mean age 15 years,SD ± 4.13) were included in the study.Intestinal biopsies from inflamed and non-inflamed mucosa of IBD patients and from controls with functional abdominal pain were cultured under aerobic and anaerobic conditions.The number of microbes belonging to the same group was calculated per weight of collected tissue.The mucus thickness in frozen samples was measured under a fluorescent microscope.RESULTS:The ratios of different bacterial groups in inflamed and non-inflamed mucosa of IBD patients and controls were specific for particular diseases.Streptococcus spp.were predominant in the inflamed mucosa of Crohn's disease(CD) patients(80% of all bacteria) ,and Lactobacillus spp.were predominant in ulcerative colitis patients(90%) .The differences were statistically significant(P = 0.01-0.001) .Lower number of bifidobacteria was observed in the whole IBD group.A relation was also found between clinical and endoscopic severity and decreased numbers of Lactobacillus and,to a lesser extent,of Streptococcus in biopsies from CD patients.The mucus layer in the inflamed sites was significantly thinner as compared to controls(P = 0.0033) and to non-inflamed areas in IBD patients(P = 0.031) .CONCLUSION:The significantly thinner mucosa of IBD patients showed a predominance of some aerobes specific for particular diseases,their numbers decreased in relation to higher clinical and endoscopic activity of the disease.

Ulcerative colitis as a polymicrobial infection characterized by sustained broken mucus barrier

To reduce medication for patients with ulcerative colitis(UC),we need to establish the etiology of UC.The intestinal microbiota of patients with inflammatory bowel disease(IBD)has been shown to differ from that of healthy controls and abundant data indicate that it changes in both composition and localization.Small intestinal bacterial overgrowth is significantly higher in IBD patients compared with controls.Probiotics have been investigated for their capacity to reduce the severity of UC.The luminal surfaces of the gastrointestinal tract are covered by a mucus layer.This normally acts as a barrier that does not allow bacteria to reach the epithelial cells and thus limits the direct contact between the host and the bacteria.The mucus layer in the colon comprises an inner layer that is firmly adherent to the intestinal mucosa,and an outer layer that can be washed off with minimal rinsing.Some bacteria can dissolve the protective inner mucus layer.Defects in renewal and formation of the inner mucus layer allow bacteria to reach the epithelium and have implications for the causes of colitis.In this review,important elements of UC pathology are thought to be the intestinal bacteria,gut mucus,and the mucosa-associated immune system.

Antimicrobial and hemolytic activity of fish epidermal mucus Cynoglossus arel and Arius caelatus

Objective:To study the antimicrobial,hemolytic activity and immunomodulatory activity of fish epidermal mucus and their chemical constituents from Cynoglossus arel(C.arel) and Alius caelatus(A.caelatus).Mucus plays an important role in the prevention of colonization by parasites,bacteria and fungi.Methods:Epidermal mucus was obtained from two marine fishes,lyophilized and the chemical composition of epidermal mucus was analysed by FT-IR analysis.The in vitro antimicrobial activity against human pathogens(fungi,gram positive and gram-negative bacteria) and also the hemolytic activity and immunomodulatory activity were determined.Results:Totally ten human pathogens were tested against the fish mucus.Out of the ten pathogens,five pathogens have proved to be sensitive to the mucus.Maximum zone of inhibition was observed against Vibrio cholera(V.cholera)(9 mm and 2 mm in diameter),followed by Staphylococcus aureus(S.aureus) with a inhibition zone of(6 mm and 3 mm),Streptococcus areus(S.areus)(5 mm and 4 mm),Vibrio parahemolyticus(V.parahemolyticus)(4 mm and 5 mm) respectively.Conclusions:The present investigation has revealed that positive progresses in the fish mucus extracts against human pathogens and hemolytic activity.But further efforts are required for the purification and isolation of the active antimicrobial compounds in order to establish their possible applications.

Adhesion of pathogenic Vibrio alginolyticus to the gill mucus of Pseudosciaena crocea

Adhesion of Vibrio alginolyticus to the gill mucus of Pseudosciaena crocea has been investigated using [ methyl-^3 H ] thymidine as isotope tracer. The results showed that： the adhesive quantity of V. alginolyticus increased with bacterial concentrations and reached equilibrium after incubated for 180 min; the higher adhesive quantity was obtained at 15 ~ 30 ℃ and sourish conditions; adhesion of V. alginolyticus could not achieved without Na^＋ , and Ca^2＋ played an auxiliary role in the bacterial adhesion; adhesion of V. alginolyticus was inhibited remarkably by starvation, heat treatment and periodic acid treatment; all of the eight kinds of carbohydrates investigated enhanced the adhesion of V. alginolyticus to the gill mucus of P. crocea, among them, glucose, mannose, fructose and maltose showed the specially enhanced adhesion. The results indicated that E alginolyticus could adhere to the gill mucus of P. crocea facilely in seawater, and this bacterial adhesion was influenced by environmental factors and closely related to superficial carbohydrate structures and some heat-sensitive structures.

Modulating intestinal mucus barrier for nanoparticles penetration by surfactants

Improving peroral delivery efficiency is always a persistent goal for both small-molecule and macromolecular drug development. However,intestinal mucus barrier which greatly impedes drug-loaded nanoparticles penetration is commonly overlooked. Therefore,in this study,taking fluorescent labeled PLGA(poly(lactic-co-glycolic acid)) nanoparticles as a tool,the influence of anionic and nonionic surfactants on mucus penetration ability of nanoparticles and their mucus barrier regulating ability were studied. The movement of PLGA nanoparticles in mucus was tracked by multiple particles tracking method(MPT).Alteration of mucus properties by addition of surfactants was evaluated by rheology and morphology study. Rat intestinal villus penetration study was used to further evaluate penetration enhancement of nanoparticles. The effective diffusivities of the nanoparticles in surfactants pretreated mucus were increased by 2–3 times and the mucus barrier regulating capacity was also surfactant type dependent. Sodium dodecyl sulfate(SDS) increased the complex viscosity and viscoelastic properties of mucus,but poloxamer presented a decreased trend. Tween 80 maintained the rheological property of the mucus. With the mucus barrier regulated by surfactants,the penetration of nanoparticles in intestinal villus was obviously increased. In summary,the mucus penetration ability of nanoparticles could be enhanced by altering mucus microenvironment with surfactants. Tween 80 which largely retains the original mucus rheology and morphology properties may be a promising candidate for facilitating nanoparticle penetration through the mucus barrier with good safety profile.

Enhanced digestion inhibition and mucus penetration of F127-modified self-nanoemulsions for improved oral delivery

Self-nanoemulsifying systems(SNEs) have excellent ability to improve the solubility ofpoorly water-soluble drugs(PWSD). However, SNEs are likely to be degraded in gastroin-testinal(GIT) when their surface is recognized by lipase/co-lipase enzyme complex, result-ing in rapid release and precipitation of encapsulated drugs. The precipitates are then cap-tured and removed by intestinal mucus, reducing the delivery efficacy of SNEs. Herein, theamphiphilic polymer Pluronic? F127 was incorporated into long and short-chain triglyc-erides(LCT, SCT) based SNEs to diminish the recognition and therefore minimized theirdegradation by enzymes and clearance by mucus. The SNEs were characterized in termsof particle size, zeta potential and stability. Ex vivo multiple particles tracking studies wereperformed by adding particle solution into fresh rat mucus. Cellular uptake of SNEs wereconducted by using E12 cells, the absorption and distribution in small intestine were alsostudied after oral administration in male Sprague-Dawley(SD) rats. The in vitro digestionrate of SNEs were found to be in following order SCT-SNE > SCT-F127-SNE > LCT-SNE > LCT-F127-SNE. Moreover, the LCT-F127-SNE was found to be most effective in enhancing cellularuptake, resulting in 3.5-fold, 2.1-fold and 1.7-fold higher than that of SCT-SNE, LCT-SNE andSCT-F127-SNE, respectively. After incubating the SNE with E12 cells, the LCT-F127-SNE ex-hibited the highest amount regarding both mucus penetration and cellular uptake, with anuptake amount number(via bicinchoninic acid(BCA) analysis) of 3.5-fold, 2.1-fold and 1.7-fold higher than that of SCT-SNE, LCT-SNE and SCT-F127-SNE, respectively. The in vivo results revealed that orally administered LCT-F127-SNE could significantly increase the bioavailability of Cyclosporine A(CsA), which was approximately 2.43-fold, 1.33-fold and 1.80-fold higher than that of SCT-SNE, SCT-F127-SNE and LCT-SNE, respectively. We address in this work that F127-modified SNEs have potentials to improve oral drug absorption by significantly reducing gastrointestinal enzymatic degradation and simultaneously enhancing mucus penetration.

Adhesion mechanisms of Vibrio fluvialis to skin mucus of Epinephelus awoara

Vibriofluvialis incubated in trypticase soy broth （TSB） showed stronger adhesion to the skin mucus of Epinephelus awoara than V. fluvialis grown on trypticase soy agar （TSA）, and this bacterial adhesion was assessed in terms of saturation kinetics. Treating bacteria with antibody against O-antigens resulted in significantly reduced bacterial adhesion. In the early growth stage, the adhering bacteria numbers increased with incubation time, peaked at 24 h, and then dropped sharply. Prior heat treatment of the mucus at various temperatures resulted in different effects on bacterial adhesion. No remarkable effect on bacterial adhesion was detected after treating mucus with protease K and trypsin, whereas periodic acid treatment significantly enhanced bacterial adhesion. Extracts of K fluvialis surface structures at high concentrations significantly inhibited bacterial adhesion, while relatively low concentrations enhanced adhesion. V. fluvialis had strong affinity for a high molecular weight mucus component. These results indicate that V.fluvialis adhered strongly to E. awoara mucus by specific adhesion and that this adhesion was influenced by many factors, including bacterial conditions, mucus, and the environment.

Development of a physiologically relevant dripping analytical method using simulated nasal mucus for nasal spray formulation analysis

Current methods for nasal spray formulations have been elementary evaluating the dripping characteristics of a formulation and have not assessed the behavior of the nasal formulation in the presence of varying types of mucus depending on the indication or diseased state. This research investigated the effects of nasal mucus on the dripping behavior of nasal formulations and focused on developing an improved in vitro analytical test method that is more physiologically relevant in characterizing nasal formulation dripping behavior. Method development was performed using simulated nasal mucus preparations for both healthy and diseased states as coatings for the dripping experiment representing a wide range of viscosity. Factors evaluated during development of this in vitro test method included amount of mucus, application of mucus, drying times, and compatibility of the mucus on a C18 Thin Layer Chromatography(TLC) substrate. The dripping behavior of nasal formulations containing a range of 1%Avicel to 3.5% Avicel was assessed by actuating the nasal spray on a perpendicular TLC plate coated with either healthy or diseased simulated nasal mucus. After actuation of the nasal spray, the dripping of the formulation on the coated TLC plate was measured after the plate was repositioned vertically. The method that was developed generated reproducible results on the dripping behavior of nasal formulations and provided critical information about the compatibility of the formulation with the nasal mucus for different diseased states, aiding in nasal spray formulation development and physical characterization of the nasal spray.

Cathelicidin stimulates colonic mucus synthesis by up-regulating MUC1 and MUC2 expression through a mitogen-activated protein kinase pathway

Objective Mucus forms the physical barrier along the gastrointestinal(GI)tract.It plays an important role to prevent mucosal damage and inflammation.Our previous finding showed that antibacterial peptide 'cathelicidin' increased mucus thickness and prevented inflammation in the colon.In the current study,we examined the protective mechanisms by which the peptide increased mucus synthesis in vitro.Methods Human colonic cell line(HT-29)was used to assess the stimulatory action of cathelicidin on mucus synthesis which was measured by the D-[6-3H] glucosamine incorporation assay.Results Human cathelicidin(LL-37)dose-dependently(10-40 μg·mL-1)and significantly stimulated mucus synthesis.Real-time PCR data showed that addition of LL-37 induced more than 50% increase in MUC1 and MUC2 mRNA levels.Treatment with MUC1 and MUC2 siRNAs normalized the stimulatory action of LL-37 on mucus synthesis.LL-37 also activated the phosphorylation of mitogen-activated protein(MAP)kinase in the cells.A specific inhibitor of the MAP kinase pathway,U0126,completely blocked the increase of MUC1 and MUC2 expression as well as mucus synthesis by LL-37.Conclusions Taken together LL-37 stimulates mucus synthesis through the activation of MUC1 and MUC2 expression and the MAP kinase pathway in human colonic cells.

Human sperm quality and lipid content after migration into normal ovulatory human cervical mucus containing low numbers of leukocytes

The aim of this study was to investigate whether a relationship exists between the presence of low numbers of leukocytes in normal ovulatory cervical mucus and sperm quality and lipid content after migration. The percentages of live, motile and morphologically normal spermatozoa, movement parameters assessed by computer-aided sperm analysis （CASA）, and ionophore-induced acrosome reaction measured by flow cytometry were determined before and after migration. High-performance liquid chromatography with ultraviolet detection was used to measure the sperm lipid content, including the various diacyl subspecies. The number of leukocytes found in solubilized mucus samples was counted using a haemocytometric method. Overall, the presence of leukocytes in the cervical mucus samples did not significantly influence sperm motility and morphology, sperm kinematic parameters, or the sperm content in sphingomyelin or cholesterol. In contrast, after migration, the decrease in various sperm diacyls and the level of induced acrosome reaction was significantly less pronounced in mucus samples containing ≥10^4 leukocytes than in mucus samples with no or rare leukocytes whereas the level of induced acrosome reaction was higher. The present data suggest that the low level of leukocytes found in normal ovulatory cervical mucus could influence the process of sperm lipid remodelling/capacitation.

Emerging role of colorectal mucus in gastroenterology diagnostics

Colonoscopy is currently the gold standard for diagnosis of inflammatory bowel disease(IBD)and colorectal cancer(CRC).This has the obvious drawback of being invasive as well as carrying a small risk.The most widely used non-invasive approaches include the use of faecal calprotectin in the case of IBD and fecal immunochemical test in the case of CRC.However,the necessity of stool collection limits their acceptability for some patients.Over the recent years,there has been emerging data looking at the role of non-invasively obtained colorectal mucus as a screening and diagnostic tool in IBD and CRC.It has been shown that the mucus rich material obtained by self-sampling of anal surface following defecation,can be used to measure various biomarkers that can aid in diagnosis of these conditions.

Antibacterial substance from mucus of a scleractinian coral,Symphyllia gigantea

Coral mucus covers the surface of coral and contains antibacterial substances as a first line of defense. Coral mucus not only enables the coral itself to resist disease, but also provides antibacterial agents for people. We collected mucus from a scleractinian coral （Symphyllia gigantea） at Sta. Sanya （China）, then extracted the antibacterial substances using 10% glacial acetic acid with the help of antiprotease inhibitors, and tested the antibacterial activity by a terrestrial bacterium （Staphylococcus aurevs） and a marine bacterium （Vibrio anguillarum）. The result showed that, there were antibacterial agents in the mucus, and their antibacterial activities were lost by treatment of the sample at 90 ℃ water for 10 min.

Effects of intrapulmonary percussive ventilation on airway mucus clearance: A bench model

AIM To determine the ability of intrapulmonary percussive ventilation(IPV) to promote airway clearance in spontaneously breathing patients and those on mechanical ventilation.METHODS An artificial lung was used to simulate a spontaneously breathing patient(Group 1), and was then connected to a mechanical ventilator to simulate a patient on mechanical ventilation(Group 2). An 8.5 mm endotracheal tube(ETT) connected to the test lung, simulated the patient airway. Artificial mucus was instilled into the mid-portion of the ETT. A filter was attached at both ends of the ETT to collect the mucus displaced proximally(mouth-piece filter) and distally(lung filter). The IPV machine was attached to the proximal end of the ETT and was applied for 10-min each to Group 1 and 2. After each experiment, the weight of the various circuit components were determined and compared to their dry weights to calculate the weight of the displaced mucus.RESULTS In Group 1(spontaneously breathing model), 26.8% ± 3.1% of the simulated mucus was displaced proximally, compared to 0% in Group 2(the mechanically ventilated model) with a P-value of < 0.01. In fact, 17% ± 1.5% of the mucus in Group 2 remained in the mid-portion of the ETT where it was initially instilled and 80% ± 4.2% was displaced distally back towards the lung(P < 0.01). There was an overall statistically significant amount of mucusmovement proximally towards the mouth-piece in the spontaneously breathing(SB) patient. There was also an overall statistically significant amount of mucus movement distally back towards the lung in the mechanically ventilated(MV) model. In the mechanically ventilated model, no mucus was observed to move towards the proximal/mouth piece section of the ETT. CONCLUSION This bench model suggests that IPV is associated with displacement of mucus towards the proximal mouthpiece in the SB patient, and distally in the MV model.

Effect of Xiaoqinglong decoction on mucus hypersecretion in the airways and cilia function in a murine model of asthma

Objective:Xiaoqinglong decoction (XQL) has been used in traditional Chinese medicine as a prescription for asthma treatment.We explored the effects of XQL on mucus hypersecretion and ciliophagy in the airways of mice in which asthma had been induced by ovalbumin (OVA).Methods:Thirty-six mice were sensitized by OVA injection (i.p.) on day-0 and day-14 and challenged with 1% OVA on day 18-22.Then,they were divided into three groups:model,carbocysteine and XQL.From day-15 to day-22,the XQL group was administered XQL (10 g/kg,p.o.) 1 hour before each aerosol challenge with OVA.To evaluate the effect of XQL on mucus hypersecretion,AB-PAS staining,measurement of serum levels of interleukin (IL)-13,bronchoalveolar lavage fluid (BALF) analyses,ciliophagy analyses,as well as coexpression of Light Chain 3 (LC3) and acetylated α-tubulin by immunofluorescence staining were undertaken.Results:Treatment with XQL (10 g/kg) attenuated mucus secretion in the airways,and reduced the positive areas of AB-PAS staining in histopathologic lung tissues (P <.05).IL-13 expression in serum (P <.01),OVA-induced inflammatory changes,and the number of white blood cells (P <.01) in BALF samples were also reduced.However,the effect on mucus secretion was less apparent in the carbocysteine group compared with the XQL group.XQL treatment also improved the cilia length and elicited a substantial reduction in ciliophagy and LC3 expression in the tracheal epithelium.Conclusion:XQL can attenuate cilia shortening,aid the clearance function of ciliated epithelial cells,and reduce mucus production in an OVA-induced asthma model in mice.XQL can inhibit mucus hypersecretion and could be a new type of pharmacotherapy.

Effects of irradiated Ergosan on the growth performance and mucus biological components of rainbow trout Oncorhynchus mykiss

Effects of irradiated and non-irradiated Ergosan extract(alginic acid) on rainbow trout growth performance and skin mucosal immunity were compared. Ergosan was irradiated at 30 kGy in a cobalt-60 irradiator. A total of 252 fish(128.03±9.4 g) were randomly divided into four equal groups,given the basal diet either unsupplemented with Ergosan(control group) or supplemented with crude Ergosan(5 g/kg),ethanol-extracted Ergosan(0.33 g/kg) or irradiated Ergosan(0.33 g/kg) according to this protocol: basal diet for 15 days,treatment diet for 15 days,basal diet for 10 days and treatment diet for 15 days. Highest growth performance was observed in fish fed irradiated Ergosan(P<0.05). Dietary administration of different Ergosan types did not cause any changes in mucus protein level,but improved alkaline phosphatase level and hemagglutination titer compared with the control(basal diet without Ergosan) on day 55 offeeding trial(P<0.05). Furthermore,the highest value of lysozyme activity was observed in gamma-irradiated Ergosan on day 55. In conclusion,gamma-irradiated Ergosan at 0.33 g/kg was found to improve growth performance and mucus biological components significantly in comparison with the control group(basal diet without Ergosan).

Colon mucus in colorectal neoplasia and beyond

Little was known about mammalian colon mucus(CM)until the beginning of the 21^(st) century.Since that time considerable progress has been made in basic research addressing CM structure and functions.Human CM is formed by two distinct layers composed of gel-forming glycosylated mucins that are permanently secreted by goblet cells of the colonic epithelium.The inner layer is dense and impenetrable for bacteria,whereas the loose outer layer provides a habitat for abundant commensal microbiota.Mucus barrier integrity is essential for preventing bacterial contact with the mucosal epithelium and maintaining homeostasis in the gut,but it can be impaired by a variety of factors,including CM-damaging switch of commensal bacteria to mucin glycan consumption due to dietary fiber deficiency.It is proven that impairments in CM structure and function can lead to colonic barrier deterioration that opens direct bacterial access to the epithelium.Bacteria-induced damage dysregulates epithelial proliferation and causes mucosal inflammatory responses that may expand to the loosened CM and eventually result in severe disorders,including colitis and neoplastic growth.Recently described formation of bacterial biofilms within the inner CM layer was shown to be associated with both inflammation and cancer.Although obvious gaps in our knowledge of human CM remain,its importance for the pathogenesis of major colorectal diseases,comprising inflammatory bowel disease and colorectal cancer,is already recognized.Continuing progress in CM exploration is likely to result in the development of a range of new useful clinical applications addressing colorectal disease diagnosis,prevention and therapy.

Toward an optimized strategy of using various airway mucus clearance techniques to treat critically ill COVID-19 patients

Coronavirus disease 2019(COVID-19)caused by acute respiratory syndrome coronavirus 2(SARS-Cov-2)is still threatening the human life and society throughout the world.For those critically ill patients,mechanical ventilation(MV)is essential to provide life support during treatment.However,both the virus infection and MV disrupt the balance between secretion and elimination of airway mucus and lead to mucus accumulation in the lung.Postmortem examination verified that the lungs in patients died of COVID-19 are indeed filled with sticky mucus,suggesting a great need to improve airway mucus clearance in critically ill COVID-19 patients.Therefore,it may be helpful to comprehensively review the current understanding regarding the changes of biochemical and rheological features of airway mucus associated with the disease,as well as the physiological principles and algorithm to decide airway clearance techniques suitable for the critically ill COVID-19 patients.Based on these considerations,optimized strategies may be developed to eliminate the airway mucus accumulated in the airways of critically ill COVID-19 patients.

Clinical pregnancy rate of women with unexplained infertility with or without cervical mucus aspiration before intrauterine insemination: A randomized controlled trial

Objective:To detect effect of removing cervical mucus before performing intrauterine insemination(IUI)on pregnancy rate in patients with unexplained infertility.Methods:The randomized controlled trial was conducted in Infertility Division of Department of Obstetrics and Gynaecology of a tertiary care hospital.Totally 80 patients of unexplained infertility were recruited.By computer generated block randomization in block size of 4 and 6,patients were randomly allocated at time of starting ovarian stimulation into the cervical mucus removal group or the non-removal of cervical mucus group before IUI,40 in each group.Ovarian stimulation with clomiphene 100 mg from day 2-6 of menstrual cycle along with human menopausal gonadotropin 150 IU was given alternate day starting from day 7.Follicular monitoring was done and further doses given as per response;trigger was planned when 1-3 follicle reach a diameter of>18 mm.IUI was planned after trigger.IUI was done as per the group allocated.Mucus cleaning was done in the cervical mucus removal group by aspirating mucus with IUI syringe and sterile cotton swab before IUI.The primary outcome was clinical pregnancy rate.Number of difficult IUIs and cycle cancellation due to ovarian hyperstimulation were also noted.Results:IUI was not done in 4 patients due to hyperstimulation.Pregnancies per IUI cycle occurred in 7.9%(3/38)in the cervical mucus removal group and 21.1%(8/38)in the non-removal of cervical mucus group.There was no statistically significant difference in clinical pregnancy rate between two groups(P=0.19).Conclusions:There is a trend towards a lower clinical pregnancy rate with removal of cervical mucus before IUI in women of unexplained infertility though the difference is not statistically significant.Further studies with large sample size need to be done on this intervention.