All grafted POSS nanoparticles changed from solid powders to liquid at room temperature. Polyurethane (PU) nanocomposites with POSS contents ranging from 1.75 to 9.72 wt % were prepared from these liquefied polyols-terminated POSS with polyepichlorohydrin (POSS–PECH). Transmission electron microscopy (TEM), scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS) were used to characterize the morphology of the POSS–PECH/PU nanocomposites. The results showed that the polyol-terminated POSS particles overcame the nanoagglomeration effect and evenly disperse in the polymeric matrix. The damping factor (tan δ) of resultant nanocomposites increased from 0.90 to 1.16, while the glass transition temperature decreased from 15.8 to 9.4 °C when POSS contents increased from 0 to 9.75 wt %. The gel content, tensile strength and Fourier transform infrared (FTIR) analyses demonstrated that the molecular thermal movement ability of the polyurethane (PU) matrix increased with increasing POSS hybrid content. Therefore, the improvement of the damping properties of the composites was mainly due to the friction-related losses occurring in the interface region between the nanoparticles and the matrix.In this work, a series of heptaphenyl siloxane trisilanol/polyhedral oligomeric silsesquioxane (T In recent decades, nanocomposites have been intensely investigated by academy and industry researchers in the field of new materials development. A large number of nanocomposites have been successfully developed and applied, becoming the key driving force for the rapid development of advanced materials for applications in aviation, aerospace, transportation and energy technologies ,5,6,7,8.Nanocomposites often exhibit excellent strength, modulus of elasticity and toughness due to the reinforcement effects of nanofillers. In these commodity engineering applications, traditional theories of polymer mechanics are usually effective in predicting and guiding the static elastic properties of composites.δ), which is attributed to 1) intra-molecular friction and molecular relaxation, 2) friction between the polymer chain and filler, 3) friction between adjacent filler particles, and 4) flexibility of the polymer chain. The value of tan δ is mainly determined by its viscoelastic behavior in the glass temperature (gT) transitional region. In this region, macromolecule chain segments tend to vibrate in phase with external vibrations. If numerous types of interactions exist between the polymer and filler, a broader tan δ transition region would be desirable. The higher the internal friction, the higher the tan δ value and the broader the transition region will be; thus, excellent energy dissipation (damping) performance of such composites will be observed [It is well known that the viscous properties of polymer composites can be expressed by the loss function known as ’tangent delta’  molecular chains from the surface of nanofillers can cause changes in conformational entropy. The associated mechanical energy loss is transformed into heat energy ,18,19,207-POSS particles were liquefied by polyepichlorohydrin (PECH) oligomer grafted by a graft-from method. POSS–PECH/polyurethane (PU) nanocomposites, all exhibiting homogeneous morphology of monodispersed POSS nanocores in the PU matrix were then fabricated and characterized. Their damping performance is starkly contrasting that observed in traditional silicon dioxide (SiO2), T7-POSS or T8-POSS composites. The mobility of PU chain segments increases with the content of POSS–PECH. The observed high tan δ and broad transition regions were attributed to increased internal friction and other interactions between the polymer chain and surface-modified POSS filler.In this study, solid T3-etherate) as the catalyst (laboratory grade). Castor oil was purchased from Tongliao Chemical Reagent Factory . Polyaryl polymethylene isocyanate (PAPI) was provided by Bayer . T7-POSS was received from Hybrid Plastics, CA .PECH was synthesized from ethylene glycol with epichlorohydrin (ECH) used as the monomer and boron trifluoride etherate  measurements were performed on a DMA +450 analyzer . The PU film was cut into 4 × 1 × 0.2 cm rectangular strips and tested in tensile mode over the temperature range from −40 to 60 °C. The heating rate was fixed at 3 °C/min. The frequencies were and 0.5, 1, 3 and 5 Hz respectively.−1 at a resolution of 4 cm−1; eight scans were collected per sample.FTIR spectra were recorded with a VERTEX 70 spectrometer  in the range of 4000–500 cmTEM images were recorded with a Tecnai G2 transmission electron microscope with 200 kV accelerating voltage . Specimens with a thickness of ca. 50 nm were prepared by ultra-cryomicrotomy at −35 °C using a Leica UCT microtome.SEM and EDS images were taken using a Philips XL-30 FEG . The samples were broken in liquid nitrogen, and all samples were coated with an ultrathin Au film by high-vacuum evaporation before observing the cross sections. At least three sections were observed to show a representative image.Tensile strength was determined by universal testing machine INSTRON 1121 , according to GB/T1040.1-2006. The samples  were tested at a strain rate of 100 mm/min.Gel permeation chromatography (GPC) analyses were carried out using PL-GPC-120 from Polymer Laboratories . The solvent used was DMF at 80 °C with a flow rate of 1.0 mL/min.1H-NMR and 13C-NMR analyses were conducted on a BRUKER 400 MHz NMR spectrometer  in CDCl3 solvent with tetramethylsilane as the internal standard.7-POSS transformed from the solid to liquid state after polymerization  and 13C-NMR(2e) spectra of POSS–PECH. The peaks at 3.62, 3.72 and 3.99 ppm in 13C-NMR spectrum of POSS–PECH is shown in 2–Cl) is at 43.13–45.07 ppm, δ(O–CH2) is at 68.98–71.13 ppm, δ(O–CH) is at 78.60–78.98 ppm, and δ(phenyl)is at 127.53–133.92 ppm.NMR spectra were useful for determining the material composition. The above analyses of FTIR and NMR spectra prove that the synthesized product was an oligomer of hydroxyl-terminated polyepichlorohydrin grafted to the POSS structure. The molecular weights of alternative POSS–PECH samples that were synthesized are shown in −1, and the stretching vibration peak of the C=O bond in the urethane bond appears at 1727 cm−1. The characteristic absorption peak of the ether bond C–O–C appears at approximately 1112 cm−1. These three features indicate that the characteristic functional group of polyurethane carbamate, NHCOO, was successfully formed. The absence of an absorption peak from isocyanate groups at 2270 cm−1 reflects the complete curing reaction.To investigate the chemical structure of the PUs with respect to various contents of POSS–PECH and pure-PECH, their FTIR spectra were obtained and are shown in The dispersion of POSS nanoparticles in the PU composites was observed by TEM. The results show that even when the POSS content reached 9.72 wt % of the total weight of the composites, no POSS particles were agglomerated, and all were uniformly distributed in the composite resin with a nearly identical size. The fracture surface of the composite presented in 2–PU composites. 2 is only 1.96 wt % of the total composite weight, agglomeration always occurs, and the cluster size after agglomeration is between 40 and 150 nm. It is also observed that the particle dispersion within clusters is not uniform. Consequently, the morphology of fracture surfaces in silica–PU composites  and the loss modulus (E”) of the sample under an oscillating load are monitored against time, temperature and frequency of the oscillation. These moduli change with frequency and temperature as the molecular motions within the polymer change. The ratio of E’/E” = tan Tg) because in this regime, without considering the chemical structure of the molecular chain, the following molecular chain motions simultaneously transform mechanical energy into thermal energy. When movable molecular chains are relatively short, under the action of external strain, there will be a large conformational change leading to the conformational entropy of materials. In the glassy state, the distance between chains is short and the interaction between chains is strong. Therefore, when the free chains move, the interaction force between the molecules that needs to be overcome is high. When the free chains move relative to each other, the internal friction between the molecular chains that needs to be overcome is large, and the internal friction between the molecular chain and the filler is also large.In general, the energy dissipation reaches a maximum near the glass transition temperature of the polymer (δ of PU nanocomposites is shown in δ of POSS-based PU nanocomposites increased from 0.91 to 1.16 with the increasing POSS content (an increase of 27.5%), whereas the damping temperature range (tan δ > 0.3) increased from ΔT = 34.4 to 44.0 °C, thus increasing by 28.1%. Meanwhile, the Tg and the initial temperature of the damping temperature range (T1) of the PU–POSS nanocomposites gradually decreased respectively: Tg to 9.4 °C from 15.8 °C and T1 to –6.9 °C from –1.1 °C. These results demonstrate that monodispersed nano-POSS particles effectively improved the damping properties of the composites, importantly demonstrating that the increased POSS content did not restrict the thermal mobility of molecular chains.The effect of POSS1 content on tan δ and ΔT of all POSS–PECH composites were higher than those of the unmodified bulk material. The tan δ decreased gradually with increasing molecular weight of POSS–PECH; although the ΔT was almost unchanged, the T1 of the damping temperature range decreased gradually with increasing molecular weight.The effects of POSS–PECH polyols with different molecular weights on the damping properties of nanocomposites were investigated, as shown in Tg of composites gradually decreased with increasing molecular weight of POSS–PECH. The initial Tg was 15.8 °C for pure PU–PECH and decreased to 0.6 °C for composites with POSS4 modified with PECH and a Mw = 4136; this is the highest range for the materials investigated in this work. This trend highlights the fact that the size of the interfacial region surrounding the POSS–PECH nanoparticles appears to be a key factor controlling the viscous properties of the system and thus the composite damping properties.At the same time, the T were almost independent of the R value. Only at R = 1.00 was the loss factor tan δ = 1.26, thus showing some improvement of damping properties. However, with the increase in the R value, the ΔT was shifted towards higher temperatures, that is, from (−12.3 ↔ +35.8 °C) to (−6.9 °C ↔ +37.1 °C). This is because the content of the hard segment increased slightly with the increase in R value, resulting in the ΔT moving towards a high temperature. These results also indicate that the improvement of the damping properties of POSS–PECH based nanocomposites is mainly due to the change in PECH-grafted POSS particle content.The R value is the molar ratio of isocyanate groups (–NCO) to hydroxyl groups (–OH). By adjusting the R value, the crosslinking density and structure of composites can be custom-tailored, and hence, it was anticipated that the damping properties could also be adjusted. The numerical data in δ of the polymer first increased and then decreased. The Tg gradually increased, the ΔT moved towards high temperature, and the range of tan δ > 0.3 increased slightly.The effects of different test frequencies on the damping properties of polyurethane polymers were analyzed using data sets presented in The analysis of the results presented above yields the following information:2–PU and unmodified POSS–PU composites  material to tan δ equal to 0.68 exhibited by the nano-SiO2–PU composite containing 9.74% of SiO2 nanoparticles. This loss function value equals only 50% of the damping capability of our composites, with POSS1–PU containing a similar concentration of surface-grafted POSS nanoparticles. The ΔT is almost unchanged compared with that related to the original material, but the T1 and the Tg of the SiO2-composite system both increase with the increasing amount of silica.When the POSS nanoparticles surface-grafted with pendent chains of PECH are distributed in the polymer matrix in the form of a monodispersed phase, the damping properties of the nanocomposites increase with the increase the nano-POSS content. This effect is completely opposite to the performance of traditional nanocomposites, such as nano-SiO2–PU composite systems is due to nanoparticles being dispersed in the soft matrix and acting as ‘physical crosslinks’, which limit the thermal movements of molecular chains in the soft matrix. These restrictions become stronger as the number of nanoparticles increases, and hence, the number of ‘physical crosslinks’ is increased. Consequently, any restrictions on the molecular thermal motion ability will lead to a decrease in the damping property of the composite material and an increase in T1 and Tg.A number of earlier studies ,29 have T1 and Tg is closely related to the crosslinking density, molecular weight between crosslinking points and the interaction between molecular chains. For this reason, we determined the following important properties of our materials: (i) The crosslinking density of the systems, which was tested by gel extraction  [For the amorphous POSS–PU nanocomposite systems, the change of mula (2) ,31:E = 1The analysis of data in 2 with similar content had little influence on the gel content of the system. Therefore, compared with SiO2–PU composites, the molecular chain thermal motions of the POSS–PU system increased with the increase of increasing POSS content due to the limitation of the decreased limitations of chemical and physical crosslinking decrease.The gel content of the PU composite decreased with the increase of POSS1 content. This indicated that the chemical crosslinking degree of the system was also decreasing. However, SiO2 composite system with similar content increased to 2400 KPa. The decreasing CED indicated that in the monodispersed state, even if the POSS particles content reached 9.72 wt %, it will not hinder the thermal motion of PU molecular chains.The CED of the PU composites decreased with increasing POSS1 content. The CED of the POSS–PU system decreased from 820 KPa (for an unmodified PU) to 400 KPa (9.72 wt % POSS1). However, the CED (a measure of the strength of molecular chain interactions) of the SiOT1 and Tg decreased with the increasing POSS content.The influence of gel content and CED on the thermal motion ability of the molecular chains of the POSS–PU system was consistent. That is, with the increase of POSS content, chemical and physical restrictions in the system were constantly reduced. Hence, the thermal motion ability of the PU molecular chain segment was enhanced. As a result, −1) and the C=O stretching region (1740–1710 cm−1), in which two absorption peaks are observed. The spectra of the stretching vibrations of N–H and –C=O in polymers with different POSS–PECH contents at the same molecular weight of POSS–PECH are shown in −1 and 1727 cm−1, respectively, and the peak position did not shift with the change of POSS–PECH content.The damping properties of polymeric materials arise from the internal friction caused by molecular chain motions and the formation and dissociation of intermolecular hydrogen bonds. The structural chemical characteristics of our nanocomposite systems were determined through FTIR, which identifies the hydrogen bonds in the analyzed systems. For the PUs, hydrogen bonding appeared in the N–H stretching region  and narrow ΔT because they lack extreme and continuous composition heterogeneity and thus cannot serve as damping materials in a broad temperature range. Considering the above issues, in this work we accomplished alleviating the above problems through: (i) Preparation of homogeneous monodispersed polymer nanocomposites utilizing bio-based PU resin as the matrix and (ii) significant broadening of the damping temperature range. Although polymers often exhibit good damping ability near the Tg, combined with broadening of damping temperature field. The proposed approach lays the foundation for a novel means of engineering the glass transition breadth and damping properties of elastomeric nanocomposites over a broadened range of application temperatures.We demonstrated here that POSS surface-grafting with oligomeric chains of variable molecular weight yields liquefied hydroxyl-terminated polyepichlorohydrin with a POSS structure. The new organic–inorganic hybrid material enables facile preparation of mono-dispersed nanocomposite systems achieving (i) total dispersion of nanoparticles providing excellent polymer nanocomposite homogeneity for POSS contents in the range 1.75 to 9.75%, (ii) excellent damping properties achieved through improvement of viscoelastic properties of interfacial zone achieved through shielding of inorganic (POSS) core by surface-grafted oligomeric molecular chains, and (iii) significant reduction of


Antibiotic use in early life has been associated with weight gain in several populations. However, associations between chronic antibiotic use and weight among adults in the general population are unknown.The NIEHS Sister Study is a longitudinal cohort of sisters of women with breast cancer. We examined associations between chronic antibiotic use (≥ 3 months) during the fourth decade of life (30–39 years) and subsequent obesity at enrollment (mean age = 55) via logistic regression. We also examined associations between chronic antibiotic use in the 5 years and 12 months prior to enrollment and weight gain after enrollment in linear mixed models. Models were adjusted for race/ethnicity, education, urban/rural status, age, and smoking.th decade of life was associated with obesity at enrollment , and use in the 5 years prior to enrollment was associated with increased BMI change after enrollment . Use of bactericidals  during the 4th decade of life was also associated with obesity at enrollment. Associations for penicillins and bactericidals were consistent across indications for use. Bacteriostatic use in the 5 years prior to enrollment was associated with a reduction in BMI after enrollment , and tetracycline use during the 4th decade of life was associated with reduced odds of obesity at enrollment . However, these inverse associations were only present for those who reported taking antibiotics for skin purposes. Cephalosporins, macrolides, quinolones, and sulfonamides were not associated with BMI change over time.In adjusted analyses , chronic penicillin use during the 4Chronic use of antibiotics during adulthood may have long-lasting impacts on BMI. Associations may differ by antibiotic class, and confounding by indication may be important for some antibiotic classes. Helicobacter pylori t status . BMI belth decade of life) to enrollment, from enrollment to the first detailed follow-up, from enrollment to the second detailed follow-up, and from the first to the second detailed follow-up. At enrollment, we used BMI from the examiner-measured height and weight data, and at all other instances used the self-report. In sensitivity analyses, we evaluated self-reported BMI at enrollment.Percent changes in BMI were also calculated for changes from a woman’s 30s  or bacteriostatic . We collapsed indications into categories of skin issues , and non-skin issues , since this categorization was the best predictor of weight change. In analyses of individual antibiotics, we considered binary variables indicating ever-use of antibiotic classes with at least 20 exposed participants, thus, we excluded aminoglycosides (n = 13), chloramphenicols (n = 5), and other beta lactams (n = 2). We also considered antibiotic use during a woman’s 30s, antibiotic use in the year prior to enrollment, and antibiotic use in the five years preceding enrollment.th and 75th percentiles for each class. These measures are included only for those who report using antibiotics.To identify important covariates for estimating the relationship between chronic antibiotic use and weight gain, we considered a directed acyclic graph (DAG) to identify possible confounders, colliders, and mediators . Variablth decade of life might not represent a model where the exposure precedes weight gain, and also narrows the time window between last reported weight and initiation of antibiotic use. We also estimated associations between use of antibiotics during their 30s and obesity at enrollment in a logistic regression model that excluded women with obesity during their 30s.We estimated prevalence odds ratios of the associations between ever use of antibiotics and obesity at enrollment in logistic regression models. We also estimated associations between use of antibiotics during the 30s and obesity at enrollment . To assess this, we considered those who initiated antibiotic use during their 30s as exposed. Those who initiated antibiotic use outside of that timeframe were excluded. This approach addressed the concern that initiation of antibiotic use prior to their 30s might capture weight gain prior to their 30s. It also addressed the concern that initiation of antibiotic use after the 4st to the 2nd follow up. We used a linear mixed regression model with random effects for subject, fixed effects for the covariates listed previously, and unstructured covariance to perform longitudinal analyses. In addition to controlling for the standard set of covariates, we controlled for BMI at enrollment. In sensitivity analyses, we omitted this variable. Since several participants reported using multiple antibiotic classes, and since users of other antibiotics were included in the referent categories for several models, we also performed sensitivity analyses where the exposed category included only those who reported using the antibiotic class of interest, and the referent category only included those who reported no antibiotic use. Participants who reported using other antibiotic classes were excluded from these models.We also took advantage of more recently collected data to minimize recall bias of weight and antibiotic use. We estimated associations between reported antibiotic use in the 12 months prior to enrollment, and five years prior to enrollment, and percent changes in BMI from enrollment to each follow up and from the 1To account for possible confounding by indication, we separately considered associations between antibiotics taken for skin and antibiotics taken for non-skin purposes in modeling antibiotic associations with weight gain. Women who did not use antibiotics were not asked about these conditions or, if asked, asked in the same way as women who used antibiotics, and therefore we were unable to assess interactions between antibiotics and indications. Instead, we compared associations for antibiotic users who reported uses for skin, and users who reported uses for non-skin, and compared those against participants who did not report using that antibiotic.We conducted several sensitivity analyses. These included evaluating modification by race/ethnicity, and diabetes status, at an alpha of 0.10. We controlled for time between first and second follow ups in longitudinal models, and also separately evaluated whether associations between antibiotic use differed by age at initiation. To assess this, we examined childhood initiation (<15 years), early adult initiation (15–45), and menopause initiation (45–55), and associations with obesity at enrollment, restricted to participants who were over the age of 55.All statistical analyses were performed in R V3.3.1.Women in the Sister Study were predominantly white, between 40 and 60 years of age at enrollment, of normal weight, and approximately half had a college degree . Of 50,8st and 2nd follow-ups, and for both adjusted and unadjusted ORs.In adjusted analyses of the associations between ever-use of antibiotics and obesity at enrollment, ever-use of cephalosporins , penicillins , quinolones , and bactericidal antibiotics  were associated with an increased odds of obesity at enrollment . Ever-usth decade of life (30s) and obesity at enrollment resulted in slightly different findings. Use of penicillins  and bactericidals , but not cephalosporins, quinolones or the broad “any antibiotic” category, during their 30s was associated with obesity at enrollment , the entirely female and mostly white makeup of the study population, and the relatively small numbers of long-term users of antibiotics. We were unable to perform a true analysis of the interaction between illness and antibiotics, as we had no data on the relevant illnesses in non-antibiotic users. For instance, if someone reported no antibiotic use, they were not asked about possible antibiotic-related illnesses. Thus, we did not know if non-users had a skin condition or a respiratory condition, and could not evaluate the effect of antibiotics among all participants with a particular condition. Instead, we were limited to estimating associations for participants who used antibiotics for skin purposes, used for non-skin purposes, or did not use that antibiotic. The confidence intervals for most of these associations were quite wide, which limited any assessment of whether the effects were different by indication category. However, we can generally conclude that the associations for penicillins remained consistent regardless of indication, while associations for tetracyclines/bacteriostatics seemed to vary by indication.Another variable with potential misclassification is self-reported weight, particularly during the participant’s 30’s. Weight may fluctuate significantly over a decade, and it is unknown whether women reported average weight, maximum weight, or minimum weight during their 30s. However, BMI in 30s was similar for antibiotic and non-antibiotics users, suggesting that outcome misclassification is likely non-differential by exposure status. Additionally, we used self-reported weight at both follow-ups, which may be subject to some exposure misclassification, particularly among obese and underweight women. However, comparisons against examiner-measured height and weight in this population suggest that women are accurate self-reporters of their height and weight, and under-reporting for obese women is rarely >10% .Recall bias for the exposure could also be a potential problem in this study. Using an antibiotic three times a week for three months or more is an unusually long period of time for antibiotic usage. Although participants may be more likely to remember taking an antibiotic for this length of time, it is also possible that women who recall taking antibiotics for an unusual length of time, such as one month, may have incorrectly reported taking the antibiotic for the full three months. Thus, the retrospective nature of this medication assessment is subject to some exposure misclassification. Additionally, although long-term antibiotic use may be more likely to impact the microbiome than short-term doses, antibiotic doses for one to two weeks are much more common. Repeated dosings of smaller amounts do appear to bring about long-lasting changes in gut microbiomes . It is pA final limitation is the external generalizability of the study. We only studied these associations in a female population that was mostly white. Therefore, we cannot draw conclusions about the relevance of these findings to men, or to a population with a higher percentage of minorities. Although we observed minimal confounding, and no effect modification by race/ethnicity, residual confounding may be present. Minority participants in the Sister Study tend to have higher incomes and to have competed more years of schooling than the general population of minority women which could in turn affect factors related to both infection and access to healthcare.Study strengths include considering indication for use in analyses, using both retrospective and prospective longitudinal data, and examination of this study question in an adult population over a long period of time. The use of both retrospective and prospective data revealed a few notable items. One, there were no associations between antibiotic use in the past 12 months and prospective weight change. It is possible that weight does not change immediately after antibiotic exposure. Changes in satiety hormones and energy storage or harvest may not stabilize until the relevant microbial communities stabilize in response to antibiotic exposure, which may take several years. Second, associations for penicillins were consistent with both retrospective and prospective data.This potential antibiotic side effect should be further investigated, as public health impact may be high. Antibiotic prescription rates are high; in 2011, healthcare providers prescribed 842 prescriptions per 1000 persons, and antibiotic prescriptions are prescribed incorrectly for between 38% to 60% of ambulatory visits , 44. Per


Immunoglobulin (Ig) G4-related disease (IgG4-RD) is a chronic inflammatory disorder characterized by high levels of serum IgG4, swollen organs with fibrosis and abundant infiltration of IgG4-positive plasmacytes.An 82-year-old male visited our hospital for an evaluation of a pancreatic enlargement and a bilateral submandibular adenopathy. Further investigation revealed elevation of serum IgG4 and bilateral lacrimal submandibular adenopathy. We diagnosed him with IgG4-related disease (IgG4-RD) and started administration of corticosteroid (CS) therapy. Both pancreatic enlargement and adenopathy rapidly improved; however, there was a new occurrence of diffuse wall thickening of the gallbladder during CS treatment.Radiological examination revealed diffuse wall thickening of the gallbladder, and its inner layer was smooth and homogenous. These findings suggested an inflammatory change, but the possibility of malignancy could not be excluded.The patient underwent laparoscopic cholecystectomy for a pathological diagnosis.Histological examination revealed a transmural infiltration of IgG4 positive plasma cells and dense fibrosis. The patient was pathologically diagnosed with IgG4 related cholecystitis presenting as an ectopic relapse.There are 2 major types of IgG4-related cholecystitis, a diffuse wall thickening type and a mass formation type. It is sometimes difficult to differentiate IgG4-related cholecystitis with gallbladder cancer.Corticosteroid (CS) is effective for induction of remission; however, we sometimes encounter disease relapse after reduction of CS dose. We should be mindful that some patients may relapse with new organ involvements even if the primary site and serum IgG4 level are well controlled. The standard treatment of IgG4-RD is oral administration of corticosteroid (CS), but surgical resection is sometimes performed because of the difficulty in differentiating IgG4-RD from malignancy. About 20% to 30% of IgG4-RD present with a relapse during the reduction of CS, however, how and where disease relapse occurs has not yet been established.Autoimmune pancreatitis (AIP) is part of a disease called IgG4-RD that often affects multiple organs including bile ducts, salivary glands, kidney and lymph nodes. IgG4-related sclerosing cholecystitis involves the gallbladder,We report a case of ectopic relapse of IgG4-RD presenting as IgG4-related cholecystitis during oral CS treatment.1.1A written informed consent for the publication of patient data was obtained from the patient.2An 82 year-old male visited our hospital for an evaluation of a pancreatic enlargement and a bilateral submandibular adenopathy. He had no abdominal symptom complaints. CT image showed diffuse swelling of the pancreas with a capsule-like rim Fig. A and intThe patient was diagnosed as IgG4-RD because of high serum IgG4 levels and organ involvement . The patient was started on 30 mg/day of CS therapy.Bilateral lacrimal submandibular adenopathy and pancreatic enlargement improved after initial CS treatment. Serum IgG4 level decreased to 315 mg/dL at 4 weeks after CS treatment.During the tapering of CS, at a dose of 8 mg/day, abdominal CT revealed diffuse wall thickening of gallbladder, although this patient did not show any abdominal symptom. Serum IgG4 level was 299 mg/dL. Laboratory data showed that white blood cell counts and C-reactive protein levels were normal.Abdominal ultrasonography showed symmetric wall thickening with echogenic foci Fig. . InflammHistological examination of surgical specimens showed a transmural infiltration of plasma cells and diffuse fibrosis without malignant cells. Immunohistochemical staining revealed abundant IgG4 positive plasma cells  , with following keywords: “IgG4” OR “Immunoglobulin G4” OR “autoimmune”, “cholecystitis” OR “gallbladder,” 197 articles were initially identified. After screening and reviews of the reference lists, 12 cases of IgG4-related sclerosing cholecystitis have been previously reported  is limited to the mass formation type. From this perspective, a distinctive morphological finding of IgG4 related cholecystitis is needed.IgG4-related sclerosing cholecystitis involves the gallbladder.In the case of suspected IgG4 related cholecystitis, gallbladder cancer and other benign disease such as xanthogranulomatous cholecystitis and chronic cholecystitis, gallbladder adenomyomatosis should be considered in the differential diagnosis. Imaging findings obtained from this case suggest that presence of diffuse wall thickening with intact mucosal layer would be a helpful morphological finding to distinguish diffuse type IgG4 related cholecystitis from gallbladder cancer.Our case has the limitation. Histological examination of surgical specimens revealed the abundant infiltration of IgG4 positive plasma cells in the gallbladder; however, it remains unclear whether IgG4-related lesion in the gallbladder existed simultaneously at the time when the patient was diagnosed with IgG4-RD. Further investigation may elucidate whether ectopic relapse occur in deceptively normal organs with potential infiltration of IgG4 positive plasma cells or actually naïve organs.In summary, IgG4-related sclerosing cholecystitis shows various patterns in morphological examinations and sometimes mimics gallbladder cancer. A large number of patients with ectopic relapse of IgG4-RD will be required for better understanding of this manifestation of IgG4-RD.The authors would like to thank Genius Plus (genius.jp.net) for English language review.Conceptualization: Keisuke Ishigami, Masahiro Shitani, Hiroshi Nakase.Data curation: Keisuke Ishigami.Investigation: Yoshiharu Masaki, Ayako Ito, Noriyuki Akutsu, Motohisa Yamamoto, Masayo Motoya, Shigeru Sasaki, Hiroki Takahashi.Resources: Yasutoshi Kimura, Tadashi Hasegawa, Ichiro Takemasa.Supervision: Hiroshi Nakase.Writing – original draft: Keisuke Ishigami, Masahiro Shitani.Writing – review & editing: Hiroshi Nakase.


IgG4-related disease (IgG4-RD) is a systemic autoimmune disease and mixed cryoglobulinemia may be caused by autoimmune diseases. However, so far only 1 case of IgG4-RD complicated with mixed cryoglobulinemia is reported. Our case further confirms the close relationship between these 2 diseases.A 55-year-old female was admitted because of dry mouth and teeth falling off.+ plasmocyte infiltration and storiform fibrosis in the interstitium of labial gland. Type III mixed cryoglobulinemia was confirmed by positive serum cryoglobulins and no monoclonal immunoglobulin in serum and urine.The patient was diagnosed as IgG4-related sialadenitis (IgG4-RS) complicated with type III mixed cryoglobulinemia. IgG4-RS was confirmed by elevated serum IgG4 levels and diffuse IgG4After treatment with prednisone and cyclophosphamide, serum cryoglobulins rapidly turned negative with the remission of IgG4-RS.Type III mixed cryoglobulinemia can be caused by IgG4-RS, and the underlying mechanisms need to be further explored. The typical histopathologic features in involved organs are significant IgG4 positive plasmacytic infiltration, storiform fibrosis, and obliterative phlebitis.IgG4-related disease (IgG4-RD) is a systemic autoimmune disease. According to reports, more than forty different organs have been involved by IgG4-RD.pe III is constituted by polyclonal IgM with rheumatoid factor activity and polyclonal IgG. Type II and III are also referred to as mixed cryoglobulins.Cryoglobulins are immunoglobulins that precipitate at temperatures less than 37°C and redissolve after rewarming. Cryoglobulinemia refers to the presence of cryoglobulins in serum. Cryoglobulins are classified into the following 3 types: type I consists of monoclonal immunoglobulin; type II is composed of monoclonal IgM with rheumatoid factor activity and polyclonal IgG; ty Now we report another case here. The case study was approved by the Ethics Review Committee of Beijing Anzhen Hospital , and informed written consent was obtained from the patient for publication of this case report.Autoimmune disease is an important cause of mixed cryoglobulinemia, but so far only 1 case of cryoglobulinemia caused by IgG4-RD has been reported by Kimaya et al.2A 55-year-old female has felt dry mouth for several months, even needing water to help swallow food sometimes. Most of her teeth have gradually taken off over the past few years. She did not feel obvious dry eyes. One month ago urinalysis found protein (+) in her urine. She never had skin purpura, ulcer, and arthralgia. Physical examination showed normal blood pressure (109/65mmHg), only 5 teeth remaining in mouth, no palpable swelling or masses in salivary glands, and no edema in lower extremities.Laboratory tests revealed mild proteinuria (0.3 g/d) with normal urinary sediment. Serum creatinine (50 μmol/L) and blood urea nitrogen (6.5mmol/L) were both normal. Serum globulin level was markedly elevated (55.9 g/L). The levels of serum IgG (43.48 g/L) and IgG4 (29.80 g/L) were also markedly elevated .The levels of serum IgA, IgM, IgG1, IgG2, and IgG3 were all normal. The levels of serum C3 (0.52 g/L) and C4 (0.04 g/L) were significantly reduced. Rheumatoid factor level was significantly increased (147.6 IU/ml). Serum cryoglobulin test was positive . Radionuclide examination of salivary glands showed mild impairment of the uptake and excretion functions in the left parotid gland, and moderate impairment in the bilateral submandibular glands. Schirmer test and tear breakup time test of both eyes were positive.+ plasmocytes (>150/HPF) and the ratio of IgG4+/IgG+ plasmocytes was 50-70%. Storiform fibrosis was observed in interstitium , IgG, IgA and C3 (1+), C1q and FRA (−) in mesangium. Immunohistochemistry staining showed no IgG4The patient was treated with prednisone  and oral cyclophosphamide (cumulant 8.5 g). After 9 months of treatment, patient's dry mouth symptoms improved significantly. Serum globulin and IgG levels both recovered to normal. Serum IgG4 level significantly dropped to near normal (3.28 g/L). Serum C3 and C4 levels both elevated to normal. Serum cryoglobulin test became negative. Urinalysis returned to normal.3The comprehensive diagnostic criteria for IgG4-RD established by Japan College of Rheumatology in 2011 are the following:1.clinical examination shows characteristic diffuse/localized swelling or masses in single or multiple organs;2.hematological examination shows elevated serum IgG4 concentrations (≥135 mg/dL);3.histopathological examination shows:(a)marked lymphocyte and plasmocyte infiltration and fibrosis;(b)+ plasmocytes: ratio of IgG4+/ IgG+ cells >40% and >10 IgG4+ plasmocytes/HPF.infiltration of IgG4 Therefore our patient can be affirmatively diagnosed as IgG4-RS.Our patient meets the above criteria (2) and (3). Although no palpable swelling or mass of the salivary glands was found at the time of hospitalization, she felt dry mouth and the uptake and excretion functions of salivary glands were impaired. We consider that no enlargement of salivary glands might be related to the chronic transformation of salivary gland lesions and obvious interstitial fibrosis. International consensus guidance statement published in 2015 pointed out that the histopathologic and immunohistochemistry features that support the diagnosis of IgG4-RD, in the proper clinical setting, can be viewed as diagnostic.,7 Based on this, we performed serum cryoglobulin test for the patient and the result was positive. So, she also suffered from cryoglobulinemia even without the clinical manifestations of vasculitis. Her cryoglobulinemia belongs to type III mixed cryoglobulinemia because no monoclonal immunoglobulin can be found in her serum and urine. Mixed cryoglobulinemia is often caused by infections such as HCV, HIV, or HBV infection, or autoimmune diseases such as Sjogren's syndrome, systemic lupus erythematosus or rheumatoid arthritis.,7 Our patient has no evidence of infection and the autoimmune diseases mentioned above. So, we think that her type III cryoglobulinemia may be caused by IgG4-RS. After treatment with prednisone and cyclophosphamide, her serum cryoglobulins rapidly turned negative with the remission of IgG4-RS, which also suggests that the 2 diseases are closely related.Positive serum rheumatoid factor and decreased C3 and C4 levels are important clues to suggest mixed cryoglobulinemia. hypothesized that dysfunction of B lymphocytes may be an underlying pathogenic mechanism of both IgG4-RD and cryoglobulinemia, in which B cell-activating factor (BAFF) may play an important role. It has been found that serum BAFF levels in patients with IgG4-RD were significantly elevated, while glucocorticoid therapy could dramatically reduce the levels of serum BAFF and IgG4. It has also observed in a patient with Sjogren's syndrome complicated with cryoglobulinemic vasculitis and elevated serum BAFF levels that serum cryoglobulins continuously turned negative after treatment with belimumab (a BAFF blocker). So, the hypothesis put forward by Kimaya et al deserves attention and should be further studied and verified in the future.The mechanism by which IgG4-RD causes cryoglobulinemia remains unclear. In 2016, Kimaya et al,10 The renal involvement of cryoglobulinemic vasculitis manifests chiefly as membrano-proliferative glomerulonephritis with thrombus-like cryoglobulin deposits within capillary lumina.,11 Therefore, the patient's kidney disease, IgM nephropathy, is an independent disease.Finally, the kidney disease in this patient is not caused by IgG4-RD or cryoglobulinemia. The major kidney lesion of IgG4-RD is tubulointerstitial nephritis with marked IgG4+ plasmacytic infiltration and storiform fibrosis in interstitium.In summary, we report here a case of IgG4-RD complicated with type III mixed cryoglobulinemia. The underlying mechanisms leading to the coexistence of these 2 diseases need to be further explored.Data curation: Ruiyu Zhang, Zhirui Zhao, Xiaoyi Xu.Formal analysis: Guoqin Wang, Hong Cheng.Resources: Lijun Sun, Hongrui Dong, Hongliang Rui.Supervision: Guoqin Wang, Hong Cheng, Yipu Chen.Validation: Hong Cheng, Yipu Chen.Visualization: Lijun Sun, Yipu Chen.Writing – original draft: Ruiyu Zhang.Writing – review & editing: Yipu Chen.


MET gene are emerging as important driver mutations for lung cancers. To identify the common clinicopathological features of MET exon 14 skipping mutations and amplification and clarify whether the two MET gene alterations cause protein overexpression were investigated using 196 lung cancer samples of Taiwan through real time-qPCR/sequencing, fluorescence in situ hybridization, and immunohistochemistry. The two MET gene alterations are both present in low frequency, ~1%, in the studied lung cancer population of Taiwan. MET exon 14 skipping mutations were identified from two early-stage patients, who were both relatively advanced in age, and did not carry other driver mutations. One was an adenocarcinoma and the other was a rare carcinosarcoma. Three gene amplifications cases were identified. Neither of the two MET gene alterations would lead to protein overexpression; hence, direct detection in nucleic acid level would be a preferred and straightforward solution for the identification of skipping mutations. The presence of MET exon 14 mutations in minor histological types of lung cancers urge to extend screening scope of this mutation in lung cancer and treatment response evaluation in clinical trials. These would be important next steps for the success of MET target therapy in clinical practice.Somatic mutations of  Lung cancer is the top leading cause of death in human population worldwide. According to the World Health organization, more than 1.6 million people died from lung cancer in 2015 [MET gene alterations, including high copy number MET gene amplification, Y1003X, and exon 14 skipping mutations [MET amplification is around 1–11% [MET exon 14 skipping mutations is around 1–4% in lung cancers [MET gene amplification and exon 14 skipping mutations are present in all major histologic types of lung cancers, including adenocarcinoma, squamous, adenosquamous, large cell and small cell [MET gene alterations would require extensive works and special techniques, such as fluorescence in situ hybridization for gene amplification, and PCR based methods coped with specific primer or probe, or sequencing for mutations. To make identification of these patients more efficiently, correlating the MET gene alterations with its protein expression level is always extensively attempted. But whether MET exon 14 skipping mutations and gene amplification confer higher MET protein expression, or relate with other specific clinicopathological features are still controversial and require more data to conclude [MET is a widely expressed receptor tyrosine kinase involved in various cellular processes, including cell growth, proliferation, survival, migration, and differentiation . To servutations . The frend 1–11% , but cound 1–11% . MET Y10nd 1–11% . The fre cancers . Both MEall cell , 16, 18,all cell , 16, 19.conclude –22.MET mutations in the lung cancer pathogenesis and in guiding diagnosis and treatment, the prevalence and clinicopathological features of the two MET alterations are investigated using a lung cancer population of Taiwan in the current study. The strengths and weaknesses of various detection methods for the MET exon 14 skipping mutations are also discussed.In view of the importance of A total of 196 formalin fixed paraffin-embedded lung cancer tissue samples collected from 2006 to 2017 with signed informed consent were requested from the tissue bank and biobank of Chang Gung Memorial Hospital, Chiayi. The study was conducted with approval from the institutional review board of Chang Gung Memorial Hospital . For each case, 1.5-mm cores were sampled from two different tumor parts and one adjacent normal part for tissue microarray construction, and seven tissue microarray (TMA) slides were prepared for the downstream FISH and IHC analysis. The clinicopathological features of these samples are summarized in MET (NM_000245.3), and 111bp fragment within exon 4 to 5 of ACTB (NM_001101.3) were amplified using primer pair listed in the MET exon 14 skipping authentic plasmid was derived from the MET wild-type authentic plasmid by two primers  and 105 copies/μl of each MET wild-type and ACTB plasmids. Five consecutive dilutions  of MET exon 14 skipping mutation plasmids were prepared and defined as 100%, 10%, 1%, 0.1%, and 0.01% respectively.The authentic plasmid containing a 529bp fragment encompassing the entire exon 13 to 14 of shown in  located MET exon 14 skipping mutation qPCR assay are listed in MET wild-type and exon 14 skipping mutation transcripts. The reaction mixture contains 1× Qiagen QuantiNova Probe master mix , 100 nM of internal control primers, 200 nM of specific primers and TaqMan probes in a 20-μl reaction volume. Real-time PCR was performed on the Qiagen Rotor-gene Q qPCR machine with the following cycling protocol: 95°C for 5 min, 45 cycles of 95°C for 5 s, 60°C for 10 s. The threshold cycle (Ct) value used for analysis was chosen at the normalized fluorescence intensity of 0.02. The qPCR products were electrophoresed in 4% agarose gel to confirm the results.Genomic DNA and total RNA were purified from the FFPE sample using AllPrep DNA/RNA Mini kit  according to the manufacturer’s protocol. For cDNA preparation, 1μg of total RNA from each sample was reverse transcribed using SuperScript III  with random hexamer. The primer and probe sequences used in the The genomic mutations contributing to the exon 14 skipping mutations were analyzed by PCR amplification of the entire exon 14 and adjacent upstream and downstream intron regions suing two primer pairs listed in MET gene amplification, FISH was performed using the MET/CEN-7 SureFISH Probe Mix  with the Histology FISH Accessory Kit (Agilent Technologies) reagents. The TMA sections were deparaffinized, rehydrated and heat pretreated at 98°C for 10 min. After cooling to room temperature for 15 min, the slides were washed twice with diluted wash buffer. The sections were then air dried and digested with pepsin for 18 min at room temperature. The pepsin digestion was terminated by washing thrice with diluted buffer and graded ethanol dehydration. To perform FISH hybridization, the fluorescence probes were reconstituted with hybridization buffer, overlaid on the TMA slides, and covered with coverslips. Hybridization reaction was performed using the Dako Hybridizer by first heating up to 90°C for 5 min, followed by gradual cooling down to 37°C and incubated for 14–20 hr. Nonspecific binding was removed by washing thrice with diluted Stringent Wash buffer at 60°C. The sections were dehydrated, mounted in medium with DAPI (Invitrogen), rested at room temperature for 24hr in the dark, and then stored at 4°C. FISH signals were observed and counted using a fluorescence microscope , and at least 50 non-overlapping nuclei were analyzed for each sample. The MET gene amplification will be interpreted if the MET gene copy numbers exceed 5 in more than 15% of counted nuclei [For the detection of d nuclei .For detection of MET protein, the anti-total MET rabbit monoclonal antibody  was used. The staining was carried out according to the manufacturer’s protocol on the BenchMark XT platform with the Ventana OptiView detection kit. The result was interpreted by a pathologist blinded to the clinical information of these patients. Only signals of staining in the cytoplasm and membrane are interpreted as positive, and the intensity was scored according to a four-tier systems: 0, no staining; 1+, weak; 2+, moderate; and 3+, strong.MET transcript is widely expressed in cells of epithelial origin; hence, the measurement the MET exon 14 skipping mutations with wild-type transcript in a single qPCR reaction together could face competition from the more abundant wild-type counterpart. For this reason, the measurements of MET exon 14 skipping mutations and wild type transcripts were performed in two separate reactions. The primer and probe set for ACTB gene detection were embedded within each reaction to ensure the appropriate setup and performance of each qPCR reaction. All PCR amplicons were chosen as short as possible to accommodate the highly fragmented nature of RNA from FFPE samples. The amplification blot and standard curve of the MET qPCR assay were generated using plasmid authentic standard and are shown in MET exon 14 skipping reached 0.1%, with the MET wild type transcripts being 1000X more abundant, and thus employed to screen the mutations in the lung cancer patient samples.ACTB internal control in both qPCR reactions for each sample. The highly fragmented nature of cDNA from FFPE samples would likely compromise qPCR amplification efficiency; hence, the cutoff value for the positivity of the MET exon 14 skipping mutations is set to Ct value of 40. Ct values for ACTB gene qPCR ranging from 21.68 to 43.4, indicate dramatic variations in RNA qualities of these clinical samples. Among the 170 samples, MET wild-type transcripts were successfully detected in 140 samples. Their Ct values ranging from 28.71 to 37.19 indicate comparatively lower MET transcript levels than that of ACTB in tumor samples. The 30 ACTB qPCR positive and MET wild-type negative samples were mainly those with late ACTB Ct values, which reflect the much less well-preserved MET transcripts in these samples. One adenocarcinoma and one carcinosarcoma, a subtype of sarcomatoid carcinoma, were found to express MET exon 14 skipping mutation transcripts. The qPCR amplification plots and Hematoxylin and Eosin (H&E) staining images of the two cases are shown in The qPCR reactions were performed on the 196 collected lung cancer samples and the detection was successful for 170 samples based on the successful amplification of MET wild type qPCR failed and 26 ACTB and MET qPCR failed samples. The two exon 14 skipping mutation-positive samples were both found to be carrying mutations (c.3802G>C) at the splicing donor site of exon 14. No mutation was identified in the 56 MET qPCR failed samples. The prevalence rate of MET exon 14 skipping mutations was 1.0% in the studied lung cancer population.Somatic mutations leading to exon14 skipping mutation transcripts in the two qPCR positive samples were further explored by sequencing the entire exon 14 and the adjacent upstream and downstream intron regions using Sanger sequencing. The mutations were also screened within the 30 MET amplification in the 196 lung cancer samples was analyzed using tissue microarray. FISH experiments were successfully performed on all samples. Among them, three were found to have MET gene amplification , three gene amplifications and 14 protein overexpressions. The frequency of the MET exon 14 skipping mutations in the current study was around 1%, which is similar to other screening results among East Asian and Caucasian population, ranging from 1 to 4% [MET exon 14 skipping mutation cases in this report are similar to those found in previous studies, including, old age (80 and 81), and mutual exclusivity to other driver mutations  [MET exon 14 skipping mutations could preferentially exist in particular patient group. Although MET exon 14 skipping mutations have been identified across different major histological subtypes of lung cancers [MET exon 14 skipping mutation cases identified from a pulmonary carcinosarcoma [MET exon 14 skipping mutation cases from other minority histologic subtypes in clinical trials for treatment evaluation.The current study screened and characterized the clinicopathological features of  1 to 4% –28. Altht shown) , 20, 25. cancers , whetherosarcoma , demonstMET exon 14 skipping mutations, the frequency of the MET amplification, 1.5%, in this lung cancer patient group are similar to those reported in other reports, ranging from 2% to 4% [MET amplification could also appear in early stage and is independent from other driver mutations; hence, it may support tumorigenesis from early stage and poses oncogenic driver activity. However, crizotinib response only observed in the high MET amplification [MET amplification in low copies exerts its oncogenic activity and responds poorly to the MET target therapy deserve further elucidation.Like that of 2% to 4% . MET ampfication , how METMET exon 14 skipping mutations may be one of the few just been uncovered for cancers, because alternative splicing transcripts could be found quite frequently in cancers [MET exon 14 skipping mutation transcripts. The wide spectrum of skipping mutations could make the molecular diagnosis challenging, since some of these mutations could result in incomplete splicing and several arise from large fragment deletions [MET exon 14 skipping mutations even with a short amplicon design adopted. Using special tissue preservative, such as PAXgene  and UMFIX , could help solve this situation. With the comprehensive sequencing profiling has become more and more important and affordable for clinical practice, it could be envisaged that adopting data directly from sequencing results using nucleic acids from fresh/frozen cell/tissue samples would be more straightforward for cancer diagnosis and treatment guidance in the future.Skipping mutations can cause various human diseases . MET exo cancers . Althoug cancers . Currenteletions , 33. TheMET gene alterations in lung cancers frequently observed in studies, including the current one, should prompt further refinement of recruiting criteria, such as including measurement of MET phosphorylation level, in future MET targeted therapy trials using MET protein as a selection marker.Overexpression of MET protein could be frequently observed and is associated with poor prognosis in lung cancer. These observations have led to the impression that MET overexpression triggers hyperactivation of MET signaling and promotes lung cancer tumorigenesis. However, the activation of MET signaling would require more than just overexpression of the receptor alone, which presumably causes the failures of several earlier MET targeted therapy trials performed on patients with MET protein overexpression , 35. DesMET exon 14 skipping mutations could help researchers in efficient identification of the mutations in their works. Although the small sample size is an inherent limitation of the study, it could help in collecting new data at a faster rate and prompt extending our understanding of MET alterations in lung cancers, such as whether MET exon 14 skipping mutations result in MET protein overexpression or not. Our findings suggest that detection methods for identification of MET exon 14 skipping mutations should be selected cautiously in the future, since MET protein overexpression is not always observed in patients with MET exon 14 skipping mutation cases; MET protein overexpression is far more prevalent than MET exon 14 skipping mutations in lung cancers. For further clarification of the important issue, MET protein expression status in cases involving MET exon 14 skipping mutations in various studies are summarized in Currently, further accumulation of data is still required before regular use of MET target therapy for lung cancers. This would mainly rely on accurate and efficient identification of suitable patients in clinical trials. The established TaqMan-based qPCR method in the study for detecting MET exon 14 skipping mutation and gene amplification are both present in low frequencies, around 1%, in the studied lung cancer population. Due to their low frequencies and do not relate to protein overexpression, detection in nucleic acid level is more suitable for detection of the two MET gene alterations. The newly identified MET exon 14 skipping mutation from a rare carcinosarcoma case urges extending screening scope for MET exon 14 skipping mutations and further evaluation in clinical trials would be an important step for the success of MET target therapy in clinical practice.In conclusion, S1 Table(DOC)Click here for additional data file.S2 Table(DOC)Click here for additional data file.


Pharmaceutical sector of Syrian Arab Republic before the war was characterized by bold and successful development since the late 1980s. With the beginning of war in the country back in March 2011, momentum has changed significantly. Traumatism, communicable diseases related to morbidity and mortality as well as wound infections became particularly hot public health concern. This relates not only to the direct victims of military conflict but also to the displaced civilians, refugees, and ordinary citizens alike. Evolving legislative framework in Syria since 1980s tolerated dispensing of antibiotics without appropriate prescription. Such practice led to spreading of antibiotic resistance among the local bacteria frequently causing both community-acquired and nosocomial infections. Laboratory findings of resistant bacteria strains among the Syrian refugees in some European countries serve as evidence of concern spreading far beyond Middle East. Practice of self-diagnosis and self-medication with antibiotics by patients themselves and restraint to pharmacist advice is widespread. A number of recommendations is presented to stakeholders to compact antibiotic resistance after the peace is established in the country. The successful implementation of such recommendations is the way to preserve shrinking golden reserve of highly potent antibiotics as it is the last defense line against resistant bacterial strains causing severe life—threatening infections. Pharmaceutical markets worldwide exhibit great diversity in terms of prescription and dispensing patterns and value-based turn over . Their dMiddle East presents a distinctively different region of the Old World . Ninety In Syria, the main source of health financing comes from the government's budget presented by the Ministry of Health and other ministries such as the Ministry of Higher Education, the Ministry of Defense, and the Ministry of Local Administration . Public Public health services in Syria are commonly of insufficient quantity and quality . Hence, patients have to visit private clinics, where they make out-of-pocket payments. Those payments slightly increased from 59.6% in 2000 to 61% in 2008 . No natiBefore the current war began in 2011, Syria was recognized among the Arab League nations for its strong domestic pharmaceutical industry . Back inComing back to contemporary momentum, we witness a disastrous war inside the Syrian Arab Republic as a consequence of complex chain of events following the Arab Spring colored revolutions . As in sSyria is no exception to similar vulnerabilities. Threatened supplies of essential medicines is currently the case even far outside major refugee migration routes  and war If we think about the nature of modern urban warfare, we could notice a long term trend that infantry weaponry is actually being made with the purpose to make wounds instead of killing at the first place. This trend in military equipment manufacturing is purely related to industrial and strategic reasons . This saHere we face another core issue even when common broad-spectrum antibiotics are at disposal of major hospitals and day care centers throughout the country. Antibiotic resistance presents an alarming threat to antimicrobial therapy. This occurring public health concern extends far beyond Syria toward other Middle Eastern neighborhood countries and the European countries alongside major refugee evacuation routes . The rooA study on the Syrian antibiotic resistance performed by Omran and Askar at Al-Mouwasat University Hospital  demonstrIn 2016, 48 Syrian migrants arrived in Italy. Upon their arrival, they received a physical examination and were subject to microbiological surveillance by blood, rectal, pharyngeal, and nasal swabs collection. Swabs were delivered and examined in local Italian clinical pathology and microbiology laboratory. Pathological analysis showed that all the 48 migrants were negative for HBV, HCV, and HIV infections. However, a large number of unusual gram-negative bacteria species were isolated and among the isolates, different strains resistant to antibiotics were found . EuropeaIn Syria, patients are frequently self-diagnosed and self-medicated, or they seek the advice of their local pharmacists  with preIt is very easy to purchase antibiotics in Syria without prescription . A crossOver long time, in a loose regulatory setting, physicians have frequently mistakenly prescribed antibiotics as a cure to diverse communicable diseases, such as flu and common cold. It is well-known that viruses are the origin of these diseases, therefore antibiotics are ineffective –42. AntiPatients, on the other hand, who may not be aware of the side effects of such antibiotic treatment, may misuse their prescribed antibiotic by stopping the course of treatment too early, when the painful symptoms begin to relieve . They maIn 2010, a cross-sectional study was carried out on 430 randomly selected adult residents of Kalamoon in Syria using standardized questionnaire. The study found that 85% had taken antibiotic medicines in the past 4 weeks and 34% were not aware of the adverse effects of antibiotics. Only 43% (out of the 85%) were prescribed the antibiotic by a physician to treat the condition, while 57% used an old prescription or took someone else's advice. This clearly indicates that the laws that control purchasing of antibiotics are ignored .It is well-known that Syria still has the largest number of pharmaceutical companies compared to most other Arab countries. Although this branch of the economy suffered heavily due to military actions, domestic companies, despite war conditions, are capable to provide antibiotics at reasonable prices. Although antibiotics are not cheap, they are affordable to many middle-income households and patients.Syria has a national-level committee designed to address antibiotic treatment related issues, including resistance. However, it has insufficient funding, resources, and leadership and thus it cannot play a significant role in controlling prescription, dispensing, and sale patterns. Moreover, Syria does not have a national policy restricting the availability of antibiotic medicines without a prescription .In Syria, three national authorities deal with antibiotic resistance: The Central Infection Prevention and Control Committee, The Directorate of Drug Affairs, and The Department of Infection Control in Hospitals' Directorates. Despite of the presence of those national bodies, the WHO officer reported that the priority given to antimicrobial resistance had been declining, due to the current war. Syria has also national laboratories with the ability to identify resistant bacteria; however, these laboratories do not produce reports or have a monitoring or reporting system for antibiotic resistance. Moreover, Syria does not participate in the regional infection control network .The Ministry of Health in Syria, three decades ago, passed a law  that determined drugs that could be sold to people without a medical prescription and antibiotics were not included in the list of drugs [Syrian Syndicate for Pharmacists—Laws and orders that coordinate pharmacy career in Syria]. Damascus, Syrian Syndicate for Pharmacists, 1994 [In Arabic]). Another law  prevented pharmacists from reselling prescribed antibiotic to the same individual without the permission of a physician and prevented physicians from prescribing an antibiotic more than twice to treat the same infection for the same individual . However, those regulations are not clearly stated or strictly enforced .Despite the global interest of the consequences of AMR, there is no sense of urgency about the current AMR status in Syria. Antimicrobial resistanceis not anymore purely a national concern. It turns to be an international issue with financial consequences. Hence, efforts should be coordinated in a Syrian national strategic plan to control the development of AMR. This can be done through reactivating the role of existed national committee and imposing more restrictions on dispensing antibiotics without prescriptions. Any savings made from the reinforcement of prevention and control activities are cost-effective and financial deficit should not be a barrier.The establishment of antimicrobial surveillance system in Syria will be a good start. This surveillance system could benefit from the instructions of European Antimicrobial Resistance Surveillance Network (EARS-Net) reporting protocol  similar Given that the high level of AMR is the result of purchasing antibiotics without prescription , 38, it We also recommend activating the role of the national committee for the rational use of antibiotics to play its expected role as a national coordinating body responsible for enhancing the prudent use of antibiotics, similar to other countries , 57. StaThe negative economic impact of AMR involves increasing mortality rate and permanently reducing the size of population and prolonging the periods of sickness and, consequently that could reduce the labor workforce efficiency. A study by Taylor estimates the GDP loss due to AMR in the MENA countries (including Syria) to range between USD 2 billions and 159 billions per year over 40 years . Such laMJa, SA, MJu, and SM all contributed equally to the acquisition of published evidence, selection, and screening of evidence for its validity and methodological quality. All three authors have revised and contributed significantly to final manuscript for important intellectual content fulfilling all ICMJE conditions for authorship.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


This research aims to experimentally investigate the potential use of a geopolymer made from various stabilizers or byproducts , slag (SL), glass powder (GP), metakaolin (MK), marble powder (MP), bottom ash (BA), rice husk ash (RHA), silica fume (SF)) to enhance the mechanical performance of soil (clay) via a deep mixing technique. Strengths of geopolymer soilcrete specimens were determined by unconfined compressive strength (UCS) tests regarding curing times (7 to 365 days) by comparing with Portland cement (PC). In addition, ultrasonic pulse velocity (UPV) tests, the effect of molarity (8–16 M), stress-strain behavior, failure modes, and microstructure  of geopolymer specimens were examined. Compared to PC, UCS responses of geopolymer specimens yielded: (i) a decreasing trend for FA-F, GP, MK, BA, and MP + FA-F, (ii) an increasing trend for FA-C, SL, and combinations of SL  favorable with fewer proportions of stabilizers, and (iii) higher increments due to long-term curing . Despite some decrements, most UCS values were found acceptable (>0.2 MPa) for sufficient enhancement of soft clay. The UCS results were mostly confirmed by UPV performances. The geopolymer specimens were also found to present: (i) strength development for alkaline concentrations from 10 to 14 M, (ii) brittle behavior of stress–strain curves that failed in axial splitting and near axial directions, and (iii) intensity of the silica peak for strength responsibility of the dense microstructure. The findings relatively support the usage of stabilizers or byproducts in the production of geopolymers for potential use in deep mixing. Thus, this research could be a basis for further efforts in this area. Moreover, the raw materials readily available for cement production are being over-consumed and results in an unsustainable source [Weak soil such as soft clay, often utilized in construction sites, needs to be enhanced in the mechanism for sufficient bearing capacity, settlement, and permeability of foundations in many civil engineering projects such as dam construction, tunnels, highway embankments, buildings, retaining walls, and so on. ,2. Numere source . In ordee source ,9 by deee source .2SiO3) or potassium silicate (K2SiO3)) formed during the geopolymerization process [Geopolymers  have emerged as a potential alternative material to Portland cement by converting the stabilizers or industrial byproducts of aluminosilicate sources  into a value-added binder using alkaline activators  or potassium hydroxide (KOH) with sodium silicate  based geopolymers are comparable to cement and lime for the improvement (grouting process) of deep soft soils ,9. Metak2SiO3 + NaOH) to stabilizer for soil-binder specimens is taken in the range from 0.85 to 1.5 on the basis of previous experiences of deep mixing [This article aims to experimentally compare the performances of using geopolymers made with various aluminosilicate stabilizers , slag (SL), bottom ash (BA), rice husk ash (RHA), glass powder (GP), silica fume (SF), marble powder (MP), and metakaolin (MK)) for deep mixing of soil (soft clay). Unconfined compressive strength (UCS) and ultrasonic pulse velocity (UPV) (7–365 days) are measured for the specimens of treated soil  columns). The binary stabilizer combinations are also investigated. To compare the responses of stabilizers, the native cement (Portland cement)-based grout is tested as a control group. Long-term performances (>90 days) of geopolymer-stabilized soils could be worth assessing for the specimens compared to ordinary Portland cement based grout. During UCS testing, the elastic modulus (Es) is also estimated from stress–strain curves, and failure planes of specimens are examined as well. Furthermore, microstructure responses are displayed with the aid of SEM and EDX analyses to elucidate the geopolymer stabilization mechanism. When less liquid content is presented in the geopolymer at the ratio of alkali activator/stabilizer to dissolve the stabilizer, then the mixture collapses instead of flows, and the workability is unmeasurable . Hence, p mixing ,23 and lThe soil (low-plasticity clay—CL) employed for testing the deep mixing was a fine-grained soil that is classified in accordance with the unified soil classification system (USCS) . It has 2/kg), the glass waste was crushed by a crusher and then completely ground by a planetary mill (using a Los Angeles machine). The silica fume (SF) in the experiments is based on silicon dioxide  in the noncrystalline (amorphous) form conformed to the recommended specifications [2/kg) with an average diameter size of 0.1 µm  that adequately fills the voids between binder particles. The high silica content and ultra-fine powder make silica fumes a highly reactive pozzolanic material for beneficial use in geopolymers [2/kg) is obtained as a byproduct from marble stone processing and was from Gaziantep industry. The metakaolin (MK) is obtained from highly processing the kaolinite type of clay (at 700 °C), with most particle grain sizes <0.044 mm. The particle sizes of BA, RHA, GP, and MP used for experiments were below the sieve size of 0.150 mm. Some chemical, physical, and index properties of materials are presented in As for the stabilizers in the production of the geopolymer, the fly ash used included both Class F  and Class C  types in accordance with specifications . Class Fications . It has polymers ,41. The 2SiO3) were used. Potable water was used to dissolve the pure flakes of NaOH to prepare an aqueous compound with a corresponding concentration of the alkaline solution. The concentration of NaOH employed for all inclusions of soil-binder specimens in this study was mainly 12 M . This has been found to be suitable in laboratory trials and matched with previous studies [2SiO3) was in a liquid gel form that had a chemical composition of 14.7% Na2O, 29.4% SiO2, and water for the remaining percentage. To add to aluminosilicate stabilizers for geopolymer production, the alkaline solution (combination of sodium hydroxide (NaOH) and sodium silicate (Na2SiO3)) was prepared in the ratio of (Na2SiO3/NaOH) = 2.5 (by weight) by blending Na2SiO3 with NaOH, and then it was left 24 h before use [In regard to the alkaline activators used for obtaining the geopolymer together with the aluminosilicate stabilizers presented above, sodium hydroxide (NaOH) and sodium silicate  ratios  with the cement replacements  suited to previous studies ,45 as weAs implied earlier, sample preparations of soft cohesive soils are the most appropriate at water contents close to their liquid limits ,35. OtheEarlier deep mixing efforts ,44,45 weAfter uniform mixtures (soil + binder mixtures) were achieved, they were cast into a cylindrical plastic mold  in order to obtain soil-binder specimens. All mixtures were compacted in three layers with a manual steel rod in the mold. Manual compaction mainly aimed to eliminate air voids for proper compaction of specimens . The comUnconfined compressive strength (UCS) tests conducted on soilcrete (soil-binder) specimens  were performed in accordance with previous specifications ,47. A unUCS performances of soilcrete specimens regarding curing times are presented in For the geopolymer made with the stabilizers alone, it was found that FA-F produced less UCS performances in comparison with PC. From the results of FA-F, an alkaline ratio of 1.25 appeared more favorable in the production of UCS responses up to 500 (7-day), 1030 (28-day), 1935 (90-day), and 2540 kPa (365-day). The lower strength from use of FA-F could be attributed to its low calcium content (4% CaO) that could be assessed as insufficient for the reorganization of alumina and silica compounds in a more stable Si-O-Al form in order to develop strong bonds at ambient conditions. Specifically, at an early age (7 and 28 days), it could be noted that dissolution of the Si and Al compounds in the alkaline solution was very slow to start the geopolymerization condensation process . ContrarAs for the SL-based geopolymer, it was able to achieve relatively higher UCS values than the ones of FA-F, FA-C, and PC, and it even produced superior performances . For replacement effects, the UCS responses increased with increasing stabilizer proportions and alkaline ratios up to r = 1.25, but beyond that, they sharply dropped at r = 1.5. Generally, the greater the alkaline ratio (r) the greater the strength from the use of the slag-based geopolymer binder. A relatively rich calcium content (34.2% CaO) of SL as an aluminosilicate precursor in alkali activation, again, can be considered a dominating factor affecting the UCS of the soil-binder (geopolymer) specimen. It is reported  that calIn regard to GP, it can be clearly observed that short-term curing performances (7 and 28 days) were low, similar to the ones of FA-F, while long-term curing (90 and 365 days) responses performed better than the ones of FA-F, nearly reaching the good magnitudes of PC. A relatively poor alumina content (1%–1.3%) and low calcium content may be a source for the low performance at an earlier age . GeneralTreatment by MK-based geopolymers for all curing times showed low UCS responses compared to PC. The UCS values remained almost unchanged, with an increasing rate up to the dosage of 25%; beyond that rate, they markedly increased (up to 2375 kPa for 365 days). It appeared that MK offered good enhancement by a large volume of stabilizer replacement and alkaline ratio. However, it should be noted that using MK during specimen preparation showed poor workability because of the relatively high specific surface area and, thus, can be considered a difficult work for deep mixing. Despite the high reactivity due to high alumina (45.7%) and silica (50.6%) contents during alkali activation, the poor workability may lead to low strengths of metakaolin . This maAs for the combinations of stabilizer-based geopolymer grouts, it was found that the addition of MP for treatment of the FA-F-based geopolymer in the stabilizer combination MP + FA-F at 20% showed a clear reduction in UCS performance compared to those of FA-F alone. This may be attributed to the fact that a poor reaction developed between MP  and the alkaline solution due to low alumina and silica contents during geopolymerization. The low strength effects of the nonpozzolanic reaction and noncementation behavior of MP seemed to dominate, similar to some applications in concrete technology .In regard to the addition of SL for enhancement of FA-F-based geopolymer in the stabilizer combination SL + FA-F at 20%, SL was able to yield higher UCS values than the ones of FA-F alone. It was observed that at earlier ages (7 and 28 days), the UCS values of this combination performed well compared to PC. On the other hand, in the long term (90 and 365 days) the addition of SL markedly produced higher responses than PC. The enhancement of FA-F by addition of SL can be explained by enhanced silica-to-alumina and silica-to-calcium ratios in the treated combination , as simiFor the ability to use BA alone and together with SL in the stabilizer combination of BA + SL at 20% for geopolymer grout, UCS responses clearly decreased with an increasing proportion of BA. Thus, it was demonstrated that low rates of BA additions for geopolymer grout were favorable for deep mixing applications. The best UCS performances  from 7 to 365 days were obtained by the addition 15% BA . Increasing alkaline ratio led to increasing strength. From the results, it can be offered that low rates of BA additions (15%–30%) had similar performances with the ones of PC at early ages of curing (7 and 28 days), while higher responses resulted in the long-term curing period (365 days). Despite having a good pozzolanic activity with high alumina (34.3%) and high silica (39.4%) contents, the decreasing UCS with increasing BA proportions in the geopolymer grout could be explained by some physical effects (very irregular shapes with cavities and pores in fine particle sizes adsorbing water)  during t2/Al2O3 ratio of the geopolymer matrix, which caused unreactive silica, due to the slower dissolution of Si and Al compounds at ambient temperature, and adsorption of a large quantity of water [Concerning the utilization of RHA and SL in the combination RHA + SL at 20%, it was observed that RHA was applicable up to 40% addition. Beyond that rate of RHA, the consistency of geopolymer grout became stiff, which is not appropriate for grout injection. UCS performances of 7 and 28 days at alkaline ratio r = 1.25 were comparable with the ones of PC that showed similar responses, whereas most of the remaining RHA additions performed better than PC. This indicated that the RHA (10%–40%) and SL mix is beneficial for geopolymer grout for deep mixing. The best UCS responses in the combination RHA + SL at 20% for both alkaline ratios were obtained by the addition of 20% RHA . For enhancement of strength by RHA, it can be said  that theof water . Higher of water . It shou2), silica fumes in a lower amount together with SL was able to form a good geopolymerization process that led to a compact microstructure and increased strength [In regard to the use of SF and SL in the combination SF + SL at 20%, it was observed that 12.5% SF addition gave the best UCS responses . Beyond that rate , the UCS responses sharply dropped, resulting in values that slightly increased with increasing SF, similar to the values of PC. The sharp decrease in UCS response (beyond 12.5% SF) could be attributed to the high surface area of SF adsorbing water during the geopolymerization process, which led to decreased surface friction of soil particles. In addition, a high amount of inactive silica, due to a high amount of SF in the combination SF + SL at 20%, could decrease the reactivity of aluminosilicate compounds because of the slower dissolution ,61. Evenstrength .For the addition of MK to SL in the combination MK + SL at 20%, it was found that UCS performances decreased with increasing MK proportions, which mostly produced better performances than PC and MK alone. On the other hand, all UCS responses of the MK addition in combination were less than SL alone, except the one at the earlier age (7 days). As compared with MK alone, it was observed that all MK additions with SL during specimen preparation offered good workability, likely due to particle effects together with the SL combination. Thus, all MK additions in the combination with SL for geopolymer grout can be applied in practice. Compared to PC and MK alone, good UCS performances of MK in the combination may be attributed to the improvement of geopolymer structure because of the good reactivity of MK by the high contents of amorphous alumina and silica compounds. On the other hand, a lower performance, specifically by a high addition of MK in the combination, may be related to the physical effects of specific surface area .From the UCS responses criticized above, it can be emphasized that RHA, MK, and SF have not been employed for treatment of FA-F, since all they have low calcium for strength development during geopolymerization in ambient conditions. They have also not been added to FA-C since they all could potentially cause poor workability and high viscosity for geopolymer grout. As a consequence , most UCThe effects of stabilizer-based geopolymer grout on the elastic modulus (Es) of soilcrete specimens are illustrated in UPV performances of soilcrete specimens  are illustrated in For Portland cement (PC) replacement, UPV values increased with replacement rate and decreased with w/b ratio . On the effect of stabilizer replacement, FA-F led to geopolymer grouts producing less UPV values, resulting in similar magnitudes with increased stabilizer proportions and ratios of alkaline activator compared to Portland cement. In comparison with FA-F, FA-C was able to yield larger UPV values, generally in an irregular trend, with an increased stabilizer rate and alkaline ratio, but again at magnitudes smaller than PC replacements. Compared to PC and other stabilizers, SL-based geopolymers seemed to result in favorite UPV values that increased with increasing stabilizer replacements up to an alkaline ratio of 1.25. However, beyond that ratio they decreased. Similar to FA-F, geopolymers made with GP mostly produced smaller UPV values, commonly in an irregular trend of increased stabilizer rate and alkaline ratio. MK-based geopolymers with increased stabilizer rates produced better UPV values than FA-F, FA-C, and GP. In addition, they appeared comparable to the ability of PC.As for the stabilizer combinations, the addition of MP (MP + FA-F at 20%) almost showed no effect on the UPV of using FA-F. On the other hand, the SL addition (SL + FA-F at 20%) with an increased alkaline ratio was able to obtain better UPV values than using FA-F alone. For utilization by the addition of BA (BA + SL at 20%), RHA (RHA + SL at 20%), SF (SF + SL at 20%) and MK (MK + SL at 20%) in combination with SL, it was observed that UPV values mostly decreased with increasing stabilizer proportions and alkaline ratios. They offered favorite UPV values in small replacements . As a consequence of UPV results , SF addiThe effects of geopolymer molarity (8 to 16 M) and water content of soil  in strength performances are illustrated in + ions in the geopolymer matrix [As shown from molarity effects , the star matrix . Consequr matrix , an emphr matrix ,60. Fromr matrix ,68,69.In regard to the effect of the water content of deep mixing soil (clay) , UCS perStress–strain curves and failure planes of soilcrete specimens are displayed in From the results , it was The SEM pictures and EDX patterns of the raw elements of soilcrete specimens are displayed in Generally, the SEM pictures  visuallyAs shown from the EDX patterns of soilcrete specimens , the int(1)Compared to PC, UCS responses of geopolymer soilcrete specimens were low for FA-F, GP, MK, BA, and MP + FA-F specimens and were high for the specimens FA-C, SL and fewer proportions of SL combinations. Among the geopolymers, SL and SF + SL (12.5% SF) offer the most favorable UCS values, and FA-C follows. The variables affecting the UCS of geopolymer specimens are stabilizer type, proportion, alkaline ratio, and curing time. Es (elastic modulus) values estimated for geopolymer specimens offer strong correlations (R > 0.90) with UCS performances. Thus, they could be beneficial for the stiffness of soil-binder columns during design.(2)In most of the treatments with stabilizer only, increasing stabilizer proportions (10% to 25%) and alkaline ratios (r = 0.85 to r = 1.25) increases UCS performances with increasing curing time. In the combinations, increasing stabilizer proportions of BA, RHA, SF, and MK added to SL leads to a decreasing UCS.(3)Increasing curing time proportionally increases UCS for both PC and the geopolymer. However, the effect of long-term curing (90 and 365 days) on UCS of geopolymer specimens results in better performances than PC. In the majority of treatments, due to the geopolymer, UCS performances of long-term curing (365-day) markedly increase compared to those of short-term curing (7 and 28 days).(4)Most UCS performances of the geopolymer specimens can be sufficiently assessed within the performance criterion of the bearing capacity (0.2–5 MPa) recommended in previous studies. However, the effect of workability, dependent upon the affecting variables, should not be underestimated.(5)UPV performances mostly confirm the UCS results of geopolymer specimens.(6)UCS responses of geopolymer specimens increase with alkaline concentrations up to (i) 14 M for FA-F and SL, (ii) 10–12 M for FA-C, and (iii) 16 M for GP. Effects of molarity (8 to 16 M) on the geopolymer specimens  generally show strength development between 10 and 14 M. UCS increases with decreasing water content of soil (LL-5 = 37 to LL-20 = 22).(7)Stress–strain curves of geopolymer soilcrete specimens (28 days) show brittle behavior, which develop shear failure planes mostly in the axial splitting and near axial orientations. The behavior and failure modes are not found to be systematically dependent on the stabilizer type, stabilizer proportion, nor alkaline ratio of the geopolymer. Following completion of the UCS test, the geopolymer specimens remain sufficiently stable without fully disintegrating. This could be beneficial to assess the damage potential of soilcrete columns for post-failure responses.(8)From microstructure analyses (UCS specimens at 28 days), the intensity of the silica peak is identified in all geopolymer specimens. No alumina and calcium peaks are found. The peak of silica appears to be in good agreement with the strength development, specifically for the geopolymer specimens with combined stabilizers .This study experimentally uncovers the potential use of geopolymer grouts made with various stabilizers or byproducts , slag (SL), glass powder (GP), marble powder (MP), bottom ash (BA), rice husk ash (RHA), silica fume (SF), and metakaolin (MK)) for ground enhancement of soil (clay) with the use of the deep mixing method. Based on the findings obtained from unconfined compressive strength (UCS) tests, ultrasonic pulse velocity (UPV) tests, stress–strain behaviors with failure planes, and microstructure analyses (SEM and EDX), the following concluding remarks can be proposed for the deep mixing of clay using geopolymer based grouts: