Cryptosporidium, Cystoisospora, and Giardia duodenalis are gastrointestinal protozoa parasites that cause diarrhea in various animals. However, information regarding the detection and phylogenetic characterization of gastrointestinal protozoa parasites in cats is limited throughout South Korea. Therefore, this study aimed to determine the detection and identify subspecies of gastrointestinal protozoa parasites in cats from South Korea. Cryptosporidium, Cystoisospora, and G. duodenalis were identified by PCR. All positive samples were subtyped by PCR and sequencing of gp60, ITS-1, tpi, bg, and gdh. A total of 290 fecal samples were collected from stray, companion, and shelter cats in six provinces. G. duodenalis was the most prevalent, with 7.93%, followed by Cystoisospora spp. (7.24%) and Cryptosporidium spp. (4.48%). In addition, C. felis (n=10), C. parvum (n=2), C. ryanae (n=1), Cystoisospora felis (n=14), Cystoisospora suis (n=5), Cystoisospora ohioensis (n=1), Cystoisospora spp. were identified in subspecies analysis of positive samples. C. felis showed a significant association with diarrhea (7.81%) and living condition (6.04%), and Cystoisospora felis in diarreha (9.38%) according to detection. Through phylogenetic analysis of the tpi, bg, and gdh genes from 23 G. duodenalispositive samples, it was confirmed that the samples of present study belonged to assemblage A, B, C, and D. The overall detection of gastrointestinal protozoan parasitic infection was 17.93%. South Korean cats have a high rate of gastrointestinal protozoan parasites infection with cat-specific Cryptosporidium and Cystoisospora, which are associated with living conditions and diarrhea symptoms. Moreover, zoonotic and other animal-specific subtype of protozoan parasites have been detected in cat feces. Cryptosporidium, Cystoisospora, and Giardia duodenalis. Cryptosporidium is a protozoan parasite that infects the small intestine and can cause diarrhea, abdominal pain, and fever in cats , companion (n=17), and shelter cats (n=124) in six provinces. The sample collection date, sex, and location were recorded using application documents. Dead companion and stray cat bodies were submitted to the Animal and Plant Quarantine Agency  to determine the cause of death and digestive lesions and diarrhea were confirmed at this time, whereas in the case of shelter cats, fecal samples were secured with the cooperation of animal shelters in each city and province in 2022. All fecal samples were stored at 4°C until further experimentations.DNA extraction was performed according to a fecal sample-based adaptation of the Maxwell® RSC PureFood GMO Kit  developed by Promega. Briefly, 200–500 mg of fecal samples were placed into a 1.5 mL microcentrifuge tube, 500 mL of CTAB Buffer was added with 35 uL of Proteinase K, and the tubes were heated at 70°C for 30 min and 95°C for 10 min after vortexing. Maxwell® RSC Cartridge preparation and loading on the Maxwell RSC Extraction System  were performed as described by Promega. All samples were eluted in 100 uL of elution buffer. Extracted DNA was stored at -20°C until further applications.Cryptosporidium, Cystoisospora, and G. duodenalis was performed by extracting DNA with specific target genes from fecal samples using a thermocycler  . The obtained sequences were deposited in GenBank under the accession numbers OQ598555-OQ598563 for gp60 of C. felis, OQ473126, OQ473172-OQ473185, OQ534549 and OQ534551-OQ534555 for ITS-1 of Cystoisospora, OQ442978-OQ442994 for tpi of G. duodenalis, OQ442958-OQ442969 for b-giardin of G. duodenalis, and OQ442970-OQ442977 for gdh of G. duodenalis.Molecular identification of , Japan) Table 1. reports . The PCRC. felis , and tpiodenalis  to identwww.megasoftware.net) using the maximum likelihood method. Phylogenetic tree stability was assessed using a bootstrap value of 1,000 replicates. For multilocus genotyping of G. duodenalis, the DNA sequences of tpi, bg, and gdh loci were concatenated in MEGA X to form the MLG, and the reference sequences were selected according to previous studies , with 3.45% of C. felis , 0.69% of C. parvum  and 0.34% of C. ryanae . C. felis showed a statistically significant difference between fecal states and living conditions (P< 0.05). However, no statistically significant differences were observed in the infection of Cryptosporidium by region, season, and gender.The detection of each parasite infection according to region, season, sex, fecal state, and living conditions is shown in Cystoisospora spp. was 7.24% , including 4.83% of Cystoisospora felis , 1.72% of Cystoisospora suis , 0.34% of Cystoisospora ohioensis  and 0.34% of Cystoisospora spp. . The detection of Cystoisospora infection was not statistically related to region, season, gender, or living conditions. Cystoisospora felis infection was affected by fecal state (P< 0.05), as diarrhea  was higher than normal  feces.The detection of G. duodenalis was 7.93% . G. duodenalis was detected in all regions, seasons, gender, fecal states, and living conditions, although no statistical differences were observed in the detection of G. duodenalis infection.The detection of Cystoisospora felis and G. duodenalis, one with C. felis and G. duodenalis, and one with Cystoisospora felis and C. parvum (data not shown).Collectively, the results revealed gastrointestinal protozoa parasite infection was confirmed in 52 out of 290  fecal samples of cats. Among them, multiple protozoa parasites were detected in five fecal samples: three were co-infected with C. felis identified using the 18S rRNA gene were sequenced using the gp60 gene for the subtype of C. felis. Based on sequence analysis of the gp60 gene, nine of the 10 isolates were successfully sequenced, while one isolate was non-typable. Four of the sequenced isolates had high homology (98.93 to 99.46%) with the GenBank sequence accession number MH240847. Two isolates had 99.43 to 99.62% homology with MW351825, and the other two isolates had 98.17 to 98.73% homology with MH240865. One isolate showed 97.71% homology to MH240868. Phylogenetic analysis of C. felis using gp60 revealed that all nine isolates in the present study clustered with C. felis subtype XIXa , B5 (n=6), C (n=1), and D (n=3) at the bg locus, and assemblage A (n=1), A2 (n=1), B (n=5), and D (n=1) at the gdh locus. Among the 23 G. duodenalis-positive fecal samples, five were amplified at all tpi, bg, and gdh loci, and concatenated nucleotide sequences were used for MLG  infection rate in cats worldwide  with MN556343 isolated from tigers in China, which is associated with Cystoisospora suis and Cystoisospora belli , which is consistent with the results of previous studies. The detection rate of G. duodenalis was relatively high in companion cats as well as overall infection, and the possibility that it can have clinical symptoms of co-infection with other pathogens as well as G. duodenalis itself was suggested in previous studies (In this study, poridium , C. feliC. felis  and Cystra felis . Howeverra felis . Therefonfection . Previounfection . The preth Korea . However Daejeon , there w Daejeon , China ( Daejeon , Iran (K Daejeon , Greece  Daejeon , and Ita Daejeon . G. duodfections . Despite studies . HoweverCryptosporidium, Cystoisospora, and G. duodenalis, which are gastrointestinal protozoa parasites from all provinces of South Korea. Cat-specific C. felis and Cystoisospora felis were identified most frequently and were associated with living conditions and diarrhea symptoms caused by infection. Moreover, C. parvum and G. duodenalis assemblages A and B, which are zoonotic subspecies, were detected, suggesting that transmission between humans and cats is possible through environmental sharing. Unexpectedly, other species-specific C. ryanae, Cystoisospora ohioensis, and G. duodenalis assemblages C and D were detected, and further research on cat infections caused by these subspecies is required.The results of the present study showed the detection according to region, gender, diarrhea symptoms, and living conditions of Data presented in this study are available upon request from the corresponding author. Representative DNA sequences from the present study were deposited in the GenBank database under the accession numbers OQ598555-OQ598563 for gp60 of C. felis, OQ473126, OQ473172-OQ473185, OQ534549, and OQ534551-OQ534555 for ITS-1 of Cystoisospora, OQ442978-OQ442994 for tpi of G. duodenalis, OQ442958-OQ442969 for b-giardin of G. duodenalis, and OQ442970-OQ442977 for gdh of G. duodenalis.The animal studies were approved by Animal and Plant Quarantine Agency. The studies were conducted in accordance with the local legislation and institutional requirements. Written informed consent was obtained from the owners for the participation of their animals in this study.CY: Data curation, Investigation, Methodology, Software, Writing – original draft, Formal Analysis. B-YM: Conceptualization, Writing – review & editing. KL: Conceptualization, Writing – review & editing. SK: Investigation, Writing – review & editing. B-KK: Funding acquisition, Writing – review & editing. M-HH: Formal Analysis, Writing – review & editing, Data curation.


Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus known to cause two major diseases: adult T-cell leukemia/lymphoma and a progressive neuromyelopathy—tropical spastic paraparesis. Many viruses may be involved in the pathogenesis of thyroiditis; however, few studies have focused on the role of HTLV-1. We aimed to investigate the association between HTLV-1 and biological thyroid dysfunction.We included 357 patients with a positive HTLV-1 serology and thyroid-stimulating hormone assay data between 2012 and 2021 in a hospital in French Guiana; we compared the prevalence of hypothyroidism and hyperthyroidism in this group with that in an HTLV-1-negative control group (722 persons) matched for sex and age.p < 0.001).The prevalence of hypothyroidism and hyperthyroidism in patients with HTLV-1 infection was significantly higher than that in the control group (11% versus 3.2% and 11.3% versus 2.3%, respectively; Our study shows, for the first time, the association between HTLV-1 and dysthyroidism in a large sample, suggesting that thyroid function exploration should be systematically implemented in this population as this may have an impact on therapeutic management. Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus that induces the transformation and clonal expansion of T cells, mainly CD4, but also CD8 T cells; it can also infect other cells in different tissues . InfectiThe seriousness of the disease is attributable to two major types of complications. HTLV-1 infections can progress to adult T-cell leukemia/lymphoma (ATLL), a malignant proliferation of T cells with a poor prognosis and a median survival of 6 months in its acute form . The secHowever, the virus can affect different organs and cause various other symptoms. Other conditions include pulmonary alveolitis; infectious dermatitis; chronic arthropathy; and autoimmune diseases such as uveitis, polymyositis, and Sjögren syndrome .Associations between thyroiditis and viral infection have been described for retroviruses  in Graves’ disease, and for HTLV-1, enterovirus, rubella, mumps virus, HSV, EBV, and parvovirus in Hashimoto’s thyroiditis . InfectiFrench Guiana is a French territory located on the north coast of South America between Brazil and Suriname; 95% of its 83,534 km²-wide surface is covered by primary forest. HTLV-1 is endemic in this region; however, its prevalence varies among ethnic groups. In this regard, there is a marked difference in the prevalence between Maroons (descendants of runaway slaves) and Wayana Amerindians  . AnotherIn this cross-sectional study, we compared the prevalence of hyperthyroidism and hypothyroidism in HTLV-1-positive and HTLV-1-negative patients; additionally, we compared the prevalence of these conditions between groups by age and sex.Data were collected from computerized patient files and anonymized. We collected the data of patients with HTLV-1-positive serologies by ELISA (enzyme-linked immunosorbent assay) with confirmation of HTLV-1 infection by Western blot or PCR (polymerase chain reaction). Serum samples from patients were tested for the detection of HTLV-1/2 antibodies using a CMIA (chemiluminescent microparticle immunoassay) method  following the manufacturer’s instructions. Patients for whom the first screening result was positive benefitted from a confirmatory assay with a Line immune assay test . This confirmation was performed at the Cayenne Hospital laboratory between 2012 and 2021; the samples were taken from adult patients who were hospitalized in Cayenne Hospital or in the remote health centers (providing care in the most isolated areas of the Amazonian forest), or from outpatients with at least one thyroid-stimulating hormone (TSH) assay performed by the hospital laboratory. In the case of patients with confirmed HTLV-1 infections, we collected data on TSH , along with the date of sample collection. As we could not confirm the TSH values, we excluded patients for whom TSH measurement was performed in a private laboratory and documented on the computerized patient files.In cases of HTLV-1 positivity, patients are routinely tested for Epstein–Barr virus and hepatitis C. Concerning the rubeola virus, we only test pregnant women (excluded from the study) and parvovirus B19 is only tested if there are clinical symptoms. In our study, none of the patients had associated hepatitis C or EBV serology and none of the patients were tested for parvovirus B19 or rubella.Furthermore, we collected data on age, sex, and place of residence classified into savannah plateau, coastal center, eastern French Guiana, and western French Guiana, and missing information.Controls were defined as patients who were hospitalized and had both negative HTLV serology and a thyroid checkup during hospitalization. Patients were randomly selected from the laboratory databases of clinical wards unrelated to thyroid disease management from 2012 to 2021. The maximum interval between requests for HTLV serology and thyroid checkups was less than 1 month. In addition, only patients with a single thyroid checkup request during this period were selected to ensure the absence of thyroid pathology and, therefore, medical follow-up.Thus, the controls were individually matched to the cases by sex and age (1:2 for women and 1:3 for men) using the Mahalanobis optimal algorithm (XLSTAT 2022.3.1.1344).A TSH level of less than 0.4 mIU/L was defined as hyperthyroidism, and a TSH level ≥4.2 mIU/L was defined as hypothyroidism. As TSH assays were performed in most cases as a screening test and not as a diagnostic test, free thyroid hormones (T4L and T3L) were mostly unmeasured; therefore, we could not differentiate between subclinical and confirmed dysthyroidism.The inclusion criteria were as follows:- Patients over 18 years of age who underwent at least one serological analysis for HTLV-1 and one TSH measurement at the Cayenne Hospital Laboratory or a remote health center- Patients who did not decline participation after being informed by letters and posters displayed in the hospital laboratoryThe exclusion criteria were as follows 2: patie®) . A descriptive analysis of the study population and clinical data was performed. Quantitative data are expressed as means and standard deviations for continuous variables, and prevalence data are expressed as frequencies and percentages. Age was categorized as ≤30 years, 30–65 years, and >65 years. The proportions of HTLV-1-positive patients with and without hyperthyroidism and hypothyroidism were compared using crude odds ratios and the χ2 test. Furthermore, we used two unconditional multiple logistic regression models to control for age, sex, and residence location (there may be ethnic or iodine input differences among the different parts of French Guiana): one with hyperthyroidism as the dependent variable and the other with hypothyroidism as the dependent variable. Because a matched analysis does not necessarily control for confounders, an unmatched analysis was performed to control for two important confounders: age and sex. To quantify the effect size with this cross-sectional data, we computed the risk difference between HTLV-1-positive and -negative patients based on age, sex, and residence location, using inverse probability weighting. Statistical p-values <0.05 were considered significant.The statistical analysis was performed using the STATA 16 software , with an average age of 64 years. Furthermore, 54% and 5% of the patients were >65 and <30 years old, respectively. The patients were primarily from the coastal regions of French Guiana.There were significantly more HTLV-1-infected women and proportionally more patients in regions with a high prevalence of HTLV-1 seropositivity, notably western French Guiana (16.96%).p < 0.001). Similarly, p < 0.001).p < 0.001). Similarly, the prevalence of hypothyroidism was significantly higher in women with HTLV-1 infection (12.87%) than that in the controls (2.11%) (p < 0.0001). The prevalence of hypothyroidism and hyperthyroidism in men with HTLV-1 infection was 7.34% and 4.59%, respectively, whereas in the control group, it was 4.43% and 4.59%, respectively, and no significant difference was observed.In women, we found a highly significant difference in hyperthyroidism prevalence: 11.89% in the HTLV-1 group and 1.85% in the control group , although not representative of the general population, was comparable to that found in two studies, conducted in Colorado and São Paulo; it was 2.4% for subclinical hyperthyroidism and 0.7% for proven hyperthyroidism . RegardiThe first case of HTLV-1-associated autoimmune thyroiditis in patients with tropical spastic paraparesis was reported by Fukazawa et al. in 1991 . In 1992Kawai et al. and Mizokami et al. reported a 6.3% and 7.4% prevalence of HTLV-1 antibodies in patients with Hashimoto thyroiditis, respectively, both of which were significantly higher than that in the general population in their provinces  . AkamineIn a sample with a larger number of HTLV-1-infected persons as compared to that in other studies, Mine et al. showed a significant difference in the prevalence of anti-TPO and anti-TG antibodies between young men with HTLV-1 seropositivity and seronegativity, which is in agreement with a link with HTLV-1 . A positViruses other than HTLV-1 have been studied for their role in thyroid dysfunction. Most studies have focused on demonstrating a link between autoimmunity and viruses. Thus, adolescents with congenital rubella were more frequently found to have anti-TPO and anti-TG antibodies and showed a greater proportion of thyroid dysfunction than the control group . AntibodThe pathophysiological mechanism of thyroid damage by HTLV-1 is still unknown, but several hypotheses drawing from knowledge about better known viruses, such as HCV, can be put forward. There is a strong genetic influence and heritability in the development of autoimmune dysthyroidism . It has These hypothetical mechanisms could also be involved in the physiopathology of the thyroid disturbances observed with HTLV-1. However, the above hypotheses drawn from an analogy with a very different virus require specific testing. HIV, a retrovirus such as HTLV-1, has been occasionally associated with thyroid dysfunction  but does not seem to offer particular insights to explain our findings. More generally, the role of HTLV-1 infection in the context of autoimmune diseases has been increasingly reported notably for rheumatoid arthritis, systemic lupus erythematosus, and Sjögren’s syndrome. Molecular mimicry has been the main suspected mechanism; however, HTLV-1-promoted altered cytokine release is critical to tolerance rupture. Hence, HTLV-1-induced changes in regulatory CD4 T-cell molecules activity affects the homeostasis of cytokines such as IFN-γ, TNF-α, TGF-β, and IL-10, which disrupts the balance in anti-inflammatory and inflammatory responses, resulting in the loss of tolerance and the development of autoimmunity . The preFor HTLV-1, although the present results support prior studies, the detailed mechanisms remain hypothetical and require further prospective studies and meta-analyses to improve our understanding.The present study has some limitations. This cross-sectional study of a population of inpatients or outpatients presumably overestimated the baseline prevalence of dysthyroidism in the general population. The risk-difference calculations imply causal relationships that could not be proven given our study design. Nevertheless, this association has been described in different populations, a replication that suggests that this was not just a chance finding and that, in our endemic region, the impact of HTLV-1 on dysthyroidism may be substantial. Thus, it may require adaptations in the standard explorations conducted in patients with dysthyroidism, who should be tested for HTLV-1, and in those with HTLV-1 infections, who should undergo TSH measurements during follow-up. The selection of matched controls was difficult in the older age groups, which distorted the specific analyses of dysthyroidism by sex and age.The hypothesis of autoimmunity could not be explored because of the reasonably low number of antibodies found in these patients (8/69); therefore, further prospective studies should investigate dysthyroidism in detail.Our findings revealed a strong association between HTLV-1 infection and biological abnormalities in TSH levels. Further prospective studies are required to explore the autoimmunity hypothesis. In practice, given our observations and previous work, we propose the conduction of a systematic TSH assay for all patients with positive HTLV-1 serologies, initially and during their follow-up. Furthermore, we propose that patients with dysthyroidism in HTLV-1-endemic regions should be tested for HTLV-1 infection. The screening of persons with HTLV-1 for dysthyroidism is potentially important in the medium and long term because it would allow an early management and control of the broad multisystemic effects of thyroid dysfunction, notably cardiac and bone morbidities for hyperthyroidism.The original contributions presented in the study are included in the article/supplementary material. Further inquiries can be directed to the corresponding author.The studies involving human participants were reviewed and approved by Ethics Committee and the Institutional Review Board of French Guiana. The patients/participants provided their written informed consent to participate in this study.Conceptualization: JD, MD, MN, NS; Data curation: JD, MD, NH, HA JMA, KDA, AN, FS, MN, NS; Formal analysis: MN, NS; Investigation: JD, NH, JMA, AN, NS; Methodology: JD, MN, NS; Project administration: JD, MD, HA, FS, MN, NS; Resources: JD, MD, NH, HA, JMA, KDA, AN, FS, NS; Statistical analysis JD, MN, NS; Supervision: MN, NS; Validation: all authors; Visualization: all authors; Manuscript writing: JD, MD, MN, NS.


This report describes a bilateral persistent median artery (PMA) originating from the ulnar artery and terminating at different levels of the upper limb. The PMA coexisted with a bilateral bifid median nerve (MN) and two bilateral interconnections (ICs characterized with the symbol -) of the MN with the ulnar nerve (UN) (MN-UN) and a unilateral reverse IC (UN-MN). Emphasis was given to the artery’s developmental background.The PMA was identified in an 80-year-old formalin-embalmed donated male cadaver.The right-sided PMA terminated at the wrist, posterior to the palmar aponeurosis. Two neural ICs were identified: the UN joined the MN deep branch (UN-MN), at the forearm’s upper third, and the MN deep stem joined the UN palmar branch (MN-UN), at the lower third . The left-sided PMA ended in the palm giving off the 3rd and 4th proper palmar digital arteries. An incomplete superficial palmar arch was identified by the contribution of the PMA, radial, and ulnar arteries. After the MN bifurcation into superficial and deep branches, the deep branches formed a loop, that was penetrated by the PMA. The MN deep branch communicated with the UN palmar branch (MN-UN).The PMA should be evaluated as a causative factor of carpal tunnel syndrome. The modified Allen's test and the Doppler ultrasound may detect the arterial flow and the angiography may depict the vessel thrombosis in complex cases. PMA could also be a "salvage" vessel for the hand supply, in radial and ulnar artery trauma. During the first gestational trimester, the median artery typically supplies the entire hand, and after the 8rve (MN) . Rodrígurve (MN) , 13, prorve (MN) . The PMArve (MN) . Thus, trve (MN) . The PMArve (MN) , 19. TheAn 80-year-old donated male cadaver was bilaterally dissected at the upper limb. The body was donated to the Anatomy Department, through the “Anatomical Gift Program” after written informed consent. In both upper limbs, the PMA originated from the UA and coexisted with a bifid MN and an interconnection (IC) of the MN deep branch with the ulnar nerve (UN) palmar branch (Martin–Gruber IC), at the forearm upper and lower thirds. Unilaterally (right side), an IC of the UN palmar branch with the MN deep branch was identified (Marinacci IC), at the forearm upper third.On the right side, the antebrachial PMA emanating posterior to the pronator teres muscle reached an anterior position in relation to the MN superficial branch Fig. , and terOn the left side, the palmar PMA, originating 3.5 cm distally to the brachial artery bifurcation, proximal to its origin from the UA, followed an ascending course lateral and proximal to the MN division into superficial and deep branches and after crossing the MN superficial branch, descended to reach an anteromedial position at the wrist, medial to the ulnar nerve Fig. . The MN,th gestational week and yields to the UA and RA. Postnatally, the MA may persist and originate from the anterior interosseous artery or directly from the UA [The current case highlights the coexistence of a bilateral PMA of origin from the UA, and variant termination (at the hand and wrist area) with a bilateral (MN-UN) and a unilateral (UN-MN) IC. Both PMAs had an anteromedial position concerning the MN, contrariwise to Haladaj et al.’s study , in whicm the UA .In the current report, the PMA originated from the UA, bilaterally. Saenz et al.  in theirIn the current case, both PMAs of origin from the UA divided the MN in the forearm, as Saenz et al.  recordedSurgeons utilizing the forearm palmar approaches, such as the Henry approach, for the internal fixation of radius and/or ulnar fractures and or proximal and/or distal radio-ulnar joint reduction and stabilization should have in mind this area’s variants. Meticulous exploration is of utmost importance to avoid injury . FurtherThe CT walls are inflexible; hence, any thickening of the components may compress the MN and its branches. A PMA when increased to 3 mm diameter could lead to CTS, in paresthesia and pain in the area innervated by the MN , 18, 19.In addition, arteriovenous malformations may have arisen from pre-existing vessels, as Krisnamurthy et al.  identifiIn CT, the bifid MN may confuse during a procedure or may cause intraoperative nerve injury . The PMAA limitation of the current report is referred to the quality of the hand tissues, mainly regarding the poor elasticity, that affected dissection and final specimen quality. Thus, photographic documentation regarding PMA branches' distribution in hand was not provided. In addition, cutaneous from muscular branches could not be identified immunohistochemically , although the dissection was careful, and the UN branching pattern was partially identified to supply the skin .The symmetrical or asymmetrical combination of variants in the wrist area described in the current report has clinical significance. The bilateral PMA coexisted with a bilateral bifid MN, a bilateral Martin–Gruber interconnection, a unilateral incomplete SPA, and a unilateral Marinacci interconnection. The asymmetrical (of different termination levels) bilateral existence of PMA is quite rare and is often associated with neural variants that may further complicate wrist surgery. It is also of note that the PMA is the main nutrient artery of the MN, and therefore, it should be preserved. Surgeons should be aware of all these variants and assess the feasibility of safe surgery through preoperative examination.


Remimazolam is a novel benzodiazepine known for its short-acting properties. The safe use of flumazenil antagonism following remimazolam infusion remains a subject of debate. We present a case of monitored anesthesia care managed to be safe through low-dose remimazolam infusion and flumazenil antagonism. Pharmacokinetic simulations revealed that low-dose remimazolam was practically useful as one of the components of multimodal sedation/analgesia. Subsequent sedation with the readministration of remimazolam after a dose of flumazenil could also be attained. Remimazolam is a novel benzodiazepine characterized by its short-acting properties with flumazenil as an antagonist . HoweverAn 80-year-old woman  was referred to our institution due to a complaint of progressively worsening hoarseness. Over the course of 10 years, an anterior mediastinal thymoma had exhibited progression unless treated with chemoradiotherapy. In response to superior vena cava syndrome, carotid artery stenting was performed as a palliative therapy. Hoarseness, attributed to left recurrent nerve palsy, had become apparent eight months before the surgery. Prior to the procedure, the patient's self-reported level of voice handicap was 56 by the Voice Handicap Index (VHI) . The patSupplemental oxygen was provided through a nasal cannula at 3 L/min, and peripheral oxygen saturation, end-tidal carbon dioxide, and respiratory rate were continuously monitored using a Capnostream® . This was done along with monitoring the electrocardiogram, heart rate, and noninvasive blood pressure. Conscious sedation was initiated with remimazolam at 0.5 mg/kg/h and dexmedetomidine at 6 µg/kg/h. After seven minutes, the patient became unresponsive to verbal commands, and the infusion rates for remimazolam and dexmedetomidine were reduced to 0.25 mg/kg/h and 0.7 µg/kg/h, respectively. Infusion rates were adjusted to maintain a respiratory rate within the range of 10 to 20/min with normocapnia. Intravenous boluses of ketamine and fentanyl were administered to suppress responses to surgical stimuli.Upon the otolaryngologist's request for patient awakening, remimazolam infusion was terminated 143 minutes after anesthesia induction. Dexmedetomidine administration was reduced from 0.5 µg/kg/h to 0.2 µg/kg/h. The patient rapidly regained consciousness within three minutes and could follow verbal commands without agitation. She did not even refuse nasoendoscopic observation. Although the patient was able to vocalize, dexmedetomidine infusion was terminated, and 0.2 mg of flumazenil was administered 155 minutes after induction. Two minutes later, a clear conversation was facilitated.Surgical correction was completed under the patient's self-assessment of improved breathy hoarseness. Intravenous remimazolam was reintroduced at 0.25 mg/kg/h, beginning 161 minutes after induction, and maintained throughout wound closure. The patient regained consciousness thirteen minutes after remimazolam termination, and her cardiorespiratory status remained stable. A total of 28.2 mg of remimazolam and 128.8 µg of dexmedetomidine were administered. The total duration of the surgery and anesthesia was 160 minutes and 207 minutes, respectively. Figure For a detailed assessment of drug administration in this case, the remimazolam effect-site concentration (Ce), remimazolam equivalent Ce, flumazenil Ce, dexmedetomidine Ce, ketamine predicted plasma concentration (Cp), norketamine Cp, ketamine equivalent Cp, and fentanyl Ce were estimated using published pharmacokinetic model parameters -11. The This MAC case demonstrates the clinical utility of the combination of low-dose remimazolam and flumazenil. Under low-dose remimazolam-based MAC, the patient showed rapid recovery after flumazenil administration. Moreover, subsequent sedation with remimazolam successfully alleviated unpleasant memories and discomfort. An ideal MAC can flexibly adjust the patient's level of consciousness along with surgical requirements while maintaining a stable cardiorespiratory status. One of the most distinguishing features of remimazolam, compared to non-benzodiazepine intravenous sedatives such as propofol, is its susceptibility to antagonism by flumazenil.Based on the pharmacokinetic simulation, Masui indicated that remimazolam overdose during anesthesia maintenance is a significant risk factor for the reappearance of an anesthesia/sedative effect of remimazolam after flumazenil reversal . Even ifRegarding this issue, our choice of a low infusion rate for remimazolam is considered to have several clinical advantages, such as quick recovery and subsequent sedation. As shown in Figure www.tivatrainer.com; Gutta B.V., Aerdenhout, The Netherlands), since a target-controlled infusion system is currently unavailable. Additionally, cardiorespiratory changes and the patient's responses must also be carefully observed.Low-dose remimazolam-based MAC has some issues that should be addressed. First, depending on the level of surgical intervention, multimodal sedation/analgesia is often necessary . Our strThis case illustrates the feasibility of conducting MAC using a combination of remimazolam and flumazenil. Given the reappearance of the effects of remimazolam following reversal by flumazenil, low-dose remimazolam infusion with multimodal sedation/analgesia provides safe and effective MAC. Subsequent sedation after flumazenil antagonism can also be easily induced with the readministration of remimazolam.


Glass membrane feeders are used in malaria research for artificial blood feeding. This study investigates the use of Hemotek membrane feeders as a standardized alternative feeding system.Plasmodium falciparum infection intensity on mosquito guts.Hemotek feeders were compared with glass feeders by assessing mosquito feeding rate, imbibed blood meal volume and While mosquito feeding rate and blood meal volume were comparable between Hemotek and glass feeders, a loss in transmission was observed using the Hemotek feeder with a conventional collagen membrane. There was no difference in transmission between both feeders when Parafilm was used as the membrane.Hemotek feeders with a Parafilm membrane can be used as an alternative feeding system for malaria transmission research. Membrane feeding assays (MFAs) allow artificial blood feeding of mosquitoes and are fundamental tools in malaria transmission research. Membrane feeding is used for mosquito colony maintenance and for research purposes in the form of direct and standard membrane feeding assays . DMFAs are commonly used to evaluate the infectiousness of naturally infected gametocyte carriers while SMFAs are the gold standard assays to quantify the capacity of immune factors or compounds to prevent transmission of in vitro cultured gametocytes. Malaria MFAs generally use water-jacketed membrane feeders from handcrafted glass that are connected to a circulating warm water bath and covered with stretched Parafilm as a feeding membrane. The Hemotek feeding system is a viable alternative that uses electrical heating of metal feeder reservoirs and the blood is covered by a collagen or stretched Parafilm membrane for insects to feed from. Hemotek feeders are commonly used in Aedes research for colony maintenance and infection experiments with arboviruses,2 but their current use is limited in malaria research and sometimes Hemotek systems have been abandoned in favour of glass feeders after observing low Plasmodium falciparum infection rates.3 Compared with custom-made glass feeders, commercial Hemotek feeders have advantages in terms of standardization, temperature control and use as stand-alone systems with a single feeder. However, the Hemotek system has never been validated directly against glass feeders. We therefore compared both feeding systems and assessed the feeding rate of Anopheles stephensi, imbibed blood meal volume and performance in P. falciparum transmission experiments.Malaria remains a major public health problem, with 247 million cases and 619 000 deaths in 2021.An. stephensi mosquitoes (Sind-Kasur strain) were maintained at 30°C and 80% humidity with a reverse day–night cycle (12:12). P. falciparum parasites (strain NF135) were cultured in an automated incubator. Venous blood was collected in LiHep BD Vacutainer tubes . Cages containing 50 mosquitoes were exposed for 10 min to feed as described previously,4 using glass midi-feeders or the Hemotek feeding system (PS6120) with 1 ml reservoirs (FU1-1). Parafilm (M laboratory film) or Hemotek collagen (MEM5) were used as feeding membranes. The blood feeding rate was first assessed by visual inspection and unfed and partially fed mosquitoes were removed. For fully engorged mosquitoes, the blood meal volume was measured in dissected abdomens immediately after feeding using the Hemoglobin Colorimetric Assay Kit  as described previously.4 Mosquito infectivity was assessed 6–8 d post-infection in a sample of 20 mosquitoes by counting the number of established oocyst in dissected 1% mercurochrome-stained midguts. Linear and negative binomial mixed effects models were used to compare feeding performance and oocyst densities between conditions, accounting for the relatedness of observations from the same experiments.P. falciparum gametocytes, oocyst densities were significantly lower in Hemotek feeders when using the Hemotek collagen membrane (p<0.0001) . There was no statistically significant difference in oocyst density between Hemotek and glass feeders when parafilm was used as the membrane (p=0.44)  or imbibed blood meal volume  Fig. A, B. Upo01) Fig. . This re44) Fig. .These findings support the use of Hemotek feeders with Parafilm for malaria transmission research. Hemotek feeders are advantageous for small-scale experiments where only a few feeders are needed at the same time. Hemotek feeders are commercially available, do not need specialized glass craftmanship—thus removing a potential source of variation—and are less fragile. However, in an experimental setup where a large number of feeders are used concurrently, glass feeders may be preferred since they can be connected in a single chain to the same circulating system, which is more cost effective than multiple Hemotek setups.P. falciparum gametocytes to An. stephensi mosquitoes. We conclude that Hemotek feeders with a Parafilm membrane (but not a collagen membrane) can be used as an alternative feeding system for malaria transmission research.We investigated the use of Hemotek membrane feeders as a standardized alternative feeding system for feeding


Blastocystis spp., Enterocytozoon bieneusi, and Giardia duodenalis infections in Leizhou black goats in Zhanjiang City, Guangdong Province, China. The total prevalence rates of Blastocystis spp., E. bieneusi, and G. duodenalis were 33.63% (76/226), 17.7% (40/226), and 24.78% (56/226), respectively. Four Blastocystis spp. subtypes , four E. bieneusi genotypes  and two assemblages (A and E) of G. duodenalis were identified. The detection of zoonotic pathogen species, genotypes, and assemblages in Leizhou black goats implied their potential involvement in the transmission of zoonotic parasitic diseases.We report the occurrence of Blastocystis spp., Enterocytozoon bieneusi, and Giardia duodenalis are three common zoonotic intestinal parasites that cause severe diarrhea and enteric diseases. Leizhou black goats are characterized by a high reproductive rate, fast growth, and good meat quality, making them one of the pre-eminent goat breeds in China. Goats are reportedly common reservoirs of these three intestinal pathogens, but no information on their prevalence or genotypic distributions in black goats in Guangdong Province, China, is available. A total of 226 fecal samples were collected from goats in Zhanjiang city and genomic DNA was extracted from them. The presence of the three pathogens was detected using nested PCR targeting the sequences encoding SSU rRNA (Blastocystis spp.), the internal transcribed spacer of rRNA , as well as beta-giardin, glutamate dehydrogenase, and triosephosphate isomerase . All PCR products were sequenced to determine the species and genotypes of the organisms. The total prevalence rates of Blastocystis spp., E. bieneusi, and G. duodenalis were 33.63% (76/226), 17.70% (40/226), and 24.78% (56/226), respectively. Four subtypes of Blastocystis spp. were detected: ST5 (n = 6), ST10 (n = 50), ST14 (n = 14), and ST21 (n = 6). Among them, ST10 was the dominant genotype, accounting for 65.79% of strains, followed by the genotypes ST14 (18.42%), zoonotic ST5 (7.89%), and ST21 (7.89%). Four genotypes of E. bieneusi were detected: CHG3 (n = 32), CM21 (n = 4), CHG1 (n = 2), and ET-L2 (n = 2). Among these, CHG3 was the dominant genotype. Assemblage E (n = 54) and concurrent assemblages A and E (n = 2) were identified in the G. duodenalis-positive goats using multilocus genotyping. Blastocystis spp., E. bieneusi, and G. duodenalis infections were common in Leizhou black goats, all of which have zoonotic genotypes, indicating the potential risk of zoonotic transmission. Our results provide basic data for the prevention and control of these three intestinal pathogens. Further studies are required to better understand their genetic characteristics and zoonotic potential in Guangdong Province.  Blastocystis spp., Enterocytozoon bieneusi, and Giardia duodenalis are three common opportunistic intestinal parasitic protozoa with wide host ranges, which include domestic animals, wildlife, and humans [d humans ,2,3. Thed humans .Blastocystis species, ST1–ST10, ST12, ST14, ST16, and ST23 have been observed in humans [Blastocystis spp. have been identified in Artiodactyla: ST1–ST8, ST10, ST12, and ST14 [Blastocystis spp. subtypes ST1, ST3, ST4, ST5, ST7, and ST10 in goats suggests their potential transmission to humans.Among the approved subtypes (ST1–ST34) of n humans . In ASEAand ST14 ,8, and Sand ST14 . The occEnterocytozoon bieneusi is a complex species, divided into 13 different genetic groups based on the sequence of the internal transcribed spacer (ITS) region of the rRNA genes [Enterocytozoon bieneusi has an estimated overall prevalence of 17.4% in sheep and 16.3% in goats worldwide [NA genes . Groups NA genes ,12,13. Eorldwide . Analyseorldwide ,15.Giardia duodenalis can be divided into eight assemblages (A–H) according to the different genotypes. The zoonotic assemblages A and B and host-adapted assemblage E of G. duodenalis have been detected in goats, and previous reports have cited assemblage E as the dominant variant [G. duodenalis infections can occur at any age, but are most commonly observed in young and immunocompromised animals. Genotyping G. duodenalis is a valuable way to identify zoonotic assemblages, and is achieved with sequence analyses of the PCR products of the glutamate dehydrogenase (gdh), β-giardin (bg), and triosephosphate isomerase genes (tpi) with multilocus genotyping (MLG) [Giardiasis has been prevalent or has occurred in outbreaks throughout the world since the 1970s, and is listed by the World Health Organization as one of the neglected diseases that endanger human health . It is e variant . In goatng (MLG) .Capra hircus) and Chuanzhong black goat (C. hircus) are the two most widely farmed meat goat breeds in Guangdong, although the Leizhou black goat is the only landrace meat goat [Blastocystis spp., E. bieneusi, and G. duodenalis, not only cause significant production losses in goats, but also pose a potential threat to public health through the cysts or oocysts excreted in goat feces. Understanding the sources and genetic diversity of Blastocystis spp., E. bieneusi, and G. duodenalis involved in these infections is essential to understanding their pathogenicity and the development of strategies for their control, because no effective vaccines or drugs are available. However, there is currently no information on Blastocystis spp., E. bieneusi, or G. duodenalis infections in Leizhou black goats.Goats are grain-saving types of livestock that provide high-quality meat, cashmere, and milk. These animals have an important role in animal husbandry in China. The Leizhou black goat  potassium dichromate solution at 4 °C until processing (<1 week). Between October and December 2022, 226 fresh fecal samples from Leizhou black goats were collected on four farms scattered within Zhanjiang city, Guangdong Province . The sam® Stool DNA Kit , according to the instructions of the kit. The DNA samples were stored at −20 °C before PCR analysis.Each fecal sample was washed with distilled water to remove the potassium dichromate before DNA extraction. Approximately 200 mg of each fecal sample was used to extract DNA with an E.Z.N.A.Blastocystis spp. And E. bieneusi were identified based on the small subunit (SSU) rRNA gene [G. duodenalis was based on the nested PCR amplification and sequence analysis of the bg, gdh, and tpi genes , 1 μL of each primer (10 μM each), 2 μL of DNA sample, and 8.5 μL of nuclease-free deionized water. The PCR products were separated with 1.20% agarose gel electrophoresis, stained with nucleic acid dye , and observed and recorded with the Azure™ c200 Gel Image Analysis System . https://blast.ncbi.nlm.nih.gov/Blast.cgi, accessed on 15 June 2023.) was used to compare each obtained sequence with the GenBank database at the National Center for Biotechnology Information (NCBI). SSU rRNA gene sequences of Blastocystis spp., ITS gene sequences of E. bieneusi, and bg gene sequences of G. duodenalis obtained  method in MEGA 7.0  to assess their genetic relationships. Bootstrap values were calculated with 1000 replicates. All of the final positive PCR amplicons were transferred to and sequenced by Sangon Biological Engineering Technology and Service Co., Ltd. . BLAST  were calculated using the Wald method in SPSS version 27.0  to evaluate the differences in the infection rates among different age groups. Differences were considered significant at p < 0.05.A χBlastocystis spp. The prevalence of Blastocystis spp. was significantly lower in growing goats  than in reserve goats , but there was no significant difference between the growing and adult goats   were positive for  = 0.09) . E. bieneusi was 17.70% . The prevalence of E. bieneusi was significantly lower in adult goats  than in growing goats , but did not differ significantly between adult goats and reserve goats . The overall detection rate of G. duodenalis , G. duodenalis was detected in 24.78% of samples . Unlike the E. bieneusi infection rate, which was highest in growing goats, the prevalence of G. duodenalis was significantly higher in reserve goats  and growing goats  than in adult goats . Based on the PCR detection of any of the three genetic loci of Blastocystis spp., E. bieneusi, and G. duodenalis were 18.58% (42/226), 5.31% (12/226), and 7.96% (18/226), respectively. Furthermore, 11.50% (26/226) of the total goats were infected with two protozoa, with 3.54% (8/226) infected with G. duodenalis and E. bieneusi, 6.19% (14/226) with G. duodenalis and Blastocystis spp., and 1.77% (4/226) with Blastocystis spp. and E. bieneusi.Viewed from the perspective of coinfection, the overall percentage of goats infected with the three pathogens was 7.08% (16/226). The percentages of goats infected with only Blastocystis spp.-positive isolates were characterized into four subtypes: ST5 (n = 6), ST10 (n = 50), ST14 (n = 14), and ST21 (n = 6) (n = 50) was the predominant subtype, and 30 of the SSU rRNA sequences shared 97–100% homology with that of an isolate derived from dairy cattle in Malaysia (MK240481), whereas the other 20 sequences shared 85–98% homology with a Blastocystis spp. from raccoon dogs in China (MT798805). Of the 14 Blastocystis spp. samples of subtype ST14, 6 shared >85% homology with a Blastocystis spp. (ON796560) from sheep in China, and the SSU rRNA sequences of the other 8 isolates shared >95% similarity with that of MW648930 from goats in Malaysia. ST21 (n = 6) was detected in growing goats (4–18 months) and reserve goats (19–30 months), and shared >85% SSU rRNA homology with ON796563 from goats in China.Based on an SSU rRNA gene sequence analysis, the 76  (n = 6) . ST5, a E. bieneusi genotypes were identified in the goats based on their genetic divergence and their positions on the ITS-based phylogenetic tree (n = 32), CM21 (n = 4), CHG1 (n = 2), and ET-L2 (n = 2). The predominant genotype of E. bieneusi in goats was CHG3 . Based on the phylogenetic analysis of the ITS sequences determined in this study and reference sequences downloaded from GenBank, all genotypes observed in the study were categorized as group 2. The CHG3 samples (n = 32) were distributed in goats of all ages and shared 95–100% ITS homology with MH822618 from goats in China. CM21 (n = 4) was detected in reserve goats (19–30 months) and shared 99% ITS homology with KU604931 from the golden monkey in China. CHG1 (n = 2) was only detected in adult goats (>30 months) and shared 95% ITS homology with MH822617 from goats in China. In contrast, ET-L2 (n = 2) was only observed in growing goats and shared 98.6% ITS homology with MT231509 from dairy cattle in Ethiopia. In the present study, four tic tree : CHG3 . A total of 42, 42, and 22 samples were positive for G. duodenalis based on the bg, gdh, and tpi sequences, respectively. Two Giardia assemblages, E and A, were detected in these samples  shared 100% bg homology with KP687765 from humans in Spain. Thirty bg sequences of assemblage E shared 97–100% homology with that of MK452880 (n = 30) from sheep in Greece, whereas another ten bg sequences shared more than 99% homology with that of KY432834 (n = 10) from dairy cattle in China , including subtypes E34 (n = 20) and E36 (n = 22), and shared 98% gdh homology with a G. duodenalis isolate derived from Tan sheep in China . Twenty-two isolates positive for G. duodenalis at the tpi locus were identified as assemblage E and shared 98–100% homology with KT922262 from calves in Ethiopia. At the Blastocystis spp. is a common intestinal parasite that infects various animals worldwide, including humans, sheep, goats, birds, and insects [Blastocystis spp. is low, many animals act as storage hosts for its transmission. Nineteen blastocyst subtypes  have been identified in animal populations in China, and new subtypes are still being discovered in different animal populations [Blastocystis spp. in sheep and goats ranges from 0.3% to 94.7% worldwide  [Blastocystis spp.-positivity rates in sheep and goats range from 0.3% to 58.0%, and the dominant subtype is ST10. However, there are significant differences in the subtypes of Blastocystis spp. in sheep and goats from different provinces or regions in China [Blastocystis spp.-positive rate in Leizhou black goats was 33.63% in the present study, higher than in goats in most other provinces of China, except Shaanxi (58%), but including Jiangsu (24.0%), Shandong (16.7%), Inner Mongolia (10.70%), Qinghai (7.5%), Heilongjiang (5.5%), and Anhui (0.3%) [Blastocystis spp. in Leizhou black goats in Zhanjiang was also higher than in many other countries, including Malaysia (30.9%) and Liberia (10.5%) [Blastocystis spp. infection in growing and adult goats (36.36%).  insects ,25. Becaulations . Among txcluded) . In Chinin China . The Blai (0.3%) ,28. The  (10.5%) ,28. In tBlastocystis spp. were identified: ST10 (n = 50), ST14 (n = 14), ST5 (n = 6), and ST21 (n = 6). Of these, ST10 was the predominant subtype, accounting for 65.79% of infections, consistent with the results of most studies in China [n = 6), was detected in the present study. The ST21 subtype has not been detected in sheep or goats in other regions of China, except recently in Tibetan sheep and Inner Mongolian goats [Rhizomys sinensis [Callosciurus erythraeus [Hydrochoerys hydrochaeris [Clethrionomys glareolus [Blastocystis spp. has two life stages: the cyst and the trophozoite. The host often becomes infected by consuming food or drinking water contaminated with cysts. Some infected humans, especially those with mixed infections of multiple pathogens or an impaired immune system, often experience symptoms such as abdominal pain, diarrhea, and vomiting, which can be life-threatening in severe cases. Apart from zoonotic subtype ST5, it remains unclear whether any other zoonotic subtypes are present in Guangdong black goats. Because black goats potentially play a role in the transmission of Blastocystis spp. to humans [Blastocystis spp. are conducted among animal husbandry workers, water sources, and black goat populations in nearby regions. Based on genetic typing and a phylogenetic analysis of the SSU rRNA gene sequence, four subtypes of  ST5 n = , and ST2in China ,28. Intean goats ,28. We asinensis , Calloscythraeus , Hydrochochaeris , and Clelareolus . In Austlareolus . Blastoc ST5 n = , and ST2Enterocytozoon bieneusi is an emerging zoonotic intestinal pathogen that infects humans and many animal species. Many studies have identified and genotyped E. bieneusi in black goats throughout the world [E. bieneusi in goats ranges from 0 to 100% worldwide, and the overall prevalence in goats is estimated to be 16.3% [E. bieneusi in black goats in Guangdong Province, with a prevalence of 17.7% (40/226), which is lower than in Egypt  [E. bieneusi and their potential role in transmitting microsporidiosis caused by this parasite.he world ,35,36,37be 16.3% . However, 11/11) , Henan  ,39,40, C, 11/11) ,40, the , 11/11) , Yunnan , 11/11) , and Qin, 11/11) , but hig, 11/11) , Shandon, 11/11) , Jiangsu, 11/11) , Anhui  , Yunnan , 11/11) ,36, and , 11/11) . These rE. bieneusi genotypes: CHG3 (n = 32), CM21 (n = 4), CHG1 (n = 2), and ET-L2 (n = 2). CHG3 was the dominant genotype, accounting for 80% of the positive samples, consistent with the results of Wang et al. [The molecular characterization identified four g et al. . CHG3 isg et al. , sheep ag et al. , black-bg et al. , and blag et al. . Howeverg et al. , east–ceg et al. , and Yung et al. , was notg et al. . This dig et al. ,39,45. G. duodenalis is widely distributed in ruminant populations, such as goats and sheep, worldwide. The total prevalence of giardiasis in sheep and goats in China is estimated to be 7.00%, although it ranges from 0.00% to 28.93% in different provinces [G. duodenalis in Leizhou black goats in Guangdong in the present study was 24.78% (56/226), higher than in goats or sheep in Anhui , Gansu , Qinghai , Henan , Shaanxi , Xinjiang , Yunnan , Hainan , Sichuan , Ningxia , and Chongqing , but lower than Inner Mongolia  [rovinces . The pre150/584) . Many fa150/584) . For exaG. duodenalis strains infecting goats or sheep belong predominantly to assemblage E, but often occur in mixed infections of assemblages A and E, assemblages A and B, or assemblages A, B, and E [G. duodenalis infection status of pig and cattle herds in different regions of Guangdong Province were conducted by our laboratory [G. duodenalis infection was relatively common in pigs  and cattle  in different regions. Interestingly, the predominant assemblages of G. duodenalis found in cattle and pig herds are assemblage A, assemblage E, and their corresponding subtypes, whereas assemblage B was not detected in any of these livestock populations. Most parts of Guangdong Province have a subtropical monsoonal climate with abundant rainfall and a dense river network, and both assemblages (A and E), which are detected in pigs, dairy cattle and goats, possess the ability to transmit to humans and animals across species. This suggests a potential transmission cycle between workers who are in close contact with both goats and other mammals. More epidemiological research is required to better understand the spread of G. duodenalis among neighboring populations and water sources. B, and E ,49,50,51B, and E , assemblB, and E . AssemblB, and E ,54. Prelboratory ,56. ThesBlastocystis spp., E. bieneusi, and G. duodenalis in Leizhou black goats in Zhanjiang City, Guangdong Province, China. The total prevalence rates of Blastocystis spp., E. bieneusi, and G. duodenalis were 33.63% (76/226), 17.7% (40/226), and 24.78% (56/226), respectively. Four Blastocystis spp. subtypes , four E. bieneusi genotypes , and two assemblages (A and E) of G. duodenalis were detected. Our findings provided valuable data for understanding the molecular epidemiology of Blastocystis spp., E. bieneusi, and G. duodenalis in Leizhou black goats. Further research is required to understand the epidemiology and genotypic characteristics of these pathogens in animal husbandry workers and water sources in nearby regions, and the possible repercussions for public health.This study evaluated the prevalence and genotypic characteristics of


The vasculature is characterized by a thin cell layer that comprises the inner wall of all blood vessels, the continuous endothelium. Endothelial cells can also be found in the eye’s cornea. And even though cornea and vascular endothelial (VE) cells differ from each other in structure, they both function as barriers and express similar junctional proteins such as the adherens junction VE-cadherin and tight-junction member claudin-5. How these barriers are controlled to maintain the barrier and thereby its integrity is of major interest in the development of potential therapeutic targets. An important target of endothelial barrier remodeling is the actin cytoskeleton, which is centrally coordinated by Rho GTPases that are in turn regulated by Rho-regulatory proteins. In this review, we give a brief overview of how Rho-regulatory proteins themselves are spatiotemporally regulated during the process of endothelial barrier remodeling. Additionally, we propose a roadmap for the comprehensive dissection of the Rho GTPase signaling network in its entirety. •Rho-regulatory proteins orchestrate spatiotemporal Rho GTPase activity patterns.•Cell architecture and post-translational modifications guide Rho regulator dynamics.•Systems analyses enable the identification of individual Rho GTPase signaling units.•Computational modeling integrates Rho GTPase signaling components into a network. Current Opinion in Physiology 2023, 34:100676EndotheliumThis review comes from a themed issue on Jeremy Pearson and Paul C EvansEdited by Endothelium”For complete overview of the section, please refer to the article collection, “https://doi.org/10.1016/j.cophys.2023.1006762468–8673/© 2023 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).The endothelium lines the luminal side of blood vessels where it controls the passage of molecules and immune cells into tissues With the use of experimental tools such as mutant analyses and GTPase activity pulldown assays, the basic principles of Rho GTPase signaling have been well established. With respect to the endothelium, this led to the general understanding that the Rho GTPases Rac1 and Cdc42 make the endothelial barrier more restrictive through the formation of lamellipodia and filopodia, respectively, whereas RhoA increases barrier permeability by forming contraction-related stress fibers Dysregulation of Rho GTPase signaling has been linked to a variety of diseases that involve endothelial barrier dysfunction, among which are vascular pathology Actin makes up roughly 10% of the total protein in endothelial cells To safeguard a reliable endothelial barrier stability, the cortical actin ring cooperates closely with two types of cellular adhesions, namely cell–cell junctions and cell-matrix adhesion complexes  [20]. EnThe Rho family of GTPases in humans consists of 20 members that can be divided into eight subfamilies Rho GEFs activate Rho GTPases by exchanging GDP for GTP Fundamental to Rho regulator specificity lies the structural core of Rho GTPases, as this determines both the conformation of its binding sites and its subcellular localization. Rho GTPases contain a G domain, a short Rho insert region, and a C-terminal hypervariable region. The G domain is where Rho regulators interact to adjust the activity status of Rho GTPases.This domain is characterized by five conserved sequence motifs (G1–G5), of which G- domain motifs G2 and G3 resemble the switch-I and switch-II regions, respectively A general concept that is believed to underlie the spatiotemporal regulation of Rho GTPases relies on the reaction-diffusion system One feature that contributes to the intracellular dynamics of Rho regulators during endothelial barrier remodeling is the spatial distribution of cellular components, including lipid structures, FAs, and components of the actin cytoskeleton In addition to the distribution of cellular components, endothelial cell geometry affects the spatiotemporal dynamics of Rho regulators d. For exAlthough over 150 Rho regulators are involved in the process of endothelial barrier remodeling, these numbers alone do not account for the wide range of actions that 20 members of the Rho GTPase family carry out during this process. In addition to the classical concept of GTP–GDP cycling, other mechanisms contribute to the regulation of Rho regulators to accomplish this plethora of actions Beside post-translational modifications, alternative mechanisms that manipulate Rho regulator dynamics include modifications at the level of gene expression Over the last decades, molecular perturbation strategies have been used to dissect the spatiotemporal regulation of individual Rho GTPase signaling pathways One way to integrate the dynamics of individual Rho GTPase signaling components, as measured by quantitative readouts of molecular perturbation, into a complete network, is by making use of computational modeling . ComputaBoolean models are known for their low computational complexity and their ability to integrate many molecular components, which is beneficial for the simulation of crosstalk with other signaling pathways. For example, Boolean modeling created new insights into the molecular mechanisms involved in endothelial-to-mesenchymal transition (EndMT). Upon drastic changes in tissue requirements, endothelial cells may differentiate into mesenchymal cells. This process inhibits the expression of VE-cadherin and thus leads to loss of cellular junctions, a scenario that is associated with the onset of various diseases. For example, in severe cases of Fuch’s dystrophy, a degenerative disease of the corneal endothelium, EndMT contributes to the development of corneal blindness In 2020, Müller and colleagues acknowledged the need for a systems analysis to reveal how Rho regulators contextualize and spatiotemporally regulate Rho GTPase signaling Understanding how Rho GTPases orchestrate endothelial barrier remodeling is a long- standing challenge. A central question is how in this course Rho-regulatory proteins are spatiotemporally regulated. Here, we provide an overview of the features that govern Rho regulator dynamics over space and time, including endothelial cell component distribution and post-translational modifications. These features together establish the framework for a model in which Rho GEFs, GAPs, and GDIs contextualize and spatially orchestrate the diffusional flux of Rho GTPases. Moreover, we evaluated comprehensive analytical approaches that may cooperatively map Rho regulator dynamics in the process of Rho GTPase signaling during endothelial barrier remodeling.1) the identification of individual Rho GTPase signaling network components with the use of family-wide expression-based systems analyses 2) validation and further characterization of Rho GTPase signaling components through quantitative readouts of molecular perturbation experiments 3) integration of individual Rho GTPase signaling component dynamics into a complete cellular network with the use of computational modeling A major challenge ahead will be to resolve Rho GTPase signaling in its entirety. Taking into consideration the most recent insights in comprehensive analytical approaches, we propose a roadmap for the dissection of spatiotemporal Rho GTPase signaling networks, including  and multiscale modeling  will likely by inevitable to resolving the exact position of GTPase signaling networks in the development of endothelium-associated diseases.This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.Jeffrey M.A. van der Krogt: Conceptualization, Methodology, Formal analysis, Writing – original draft. Ivanka J.E. van der Meulen: Conceptualization, Writing – review & editing, Supervision. Jaap D. van Buul: Conceptualization, Writing – review & editing, Supervision, Project administration.Nothing declared.


Improved chemotherapy and radiotherapy treatment protocols fortunately increased the survival rates over the years, and more improvement is anticipated in the coming years. However, these treatments are accompanied with late effects and may result in infertility or subfertility in those survivors. Sadly, most cancer survivors realize the effects of the treatment they received on their fertility very late, when they failed attempting to have children. Thus, it is of great importance that options for fertility preservation are offered and/or made available for young cancer survivors. Therefore, health care providers should be aware of the existence of fertility-related issues in their patients and to be able to discuss the impact of the cancer and its treatment on their fertility and possible fertility preservation options. In this study, we aim to explore the perspective of oncologists and physicians dealing with and treating patients with cancer on fertility preservation. We demonstrated in this study that oncologists in Oman are supportive of fertility preservation. The lack of knowledge and unavailability of well-structured fertility preservation services in the country hinders the initiation of fertility preservation discussions. Our findings will pave the way for improving the care for cancer survivors for better quality of life.Background: Improved chemotherapy and radiotherapy treatment protocols, fortunately, increased the rates of cancer survivors over the years. However, these treatments may result in infertility or subfertility. Oncologists are considered the gateway for knowledge about cancer and its treatments’ effects. Several studies showed that many oncologists do not discuss fertility preservation with their patients. This study aimed to explore the perspective of oncologists in Oman on fertility preservation. Methods: A cross-sectional study of physicians and surgeons dealing with patients with cancer was conducted from June 2021 to December 2021. A standardized and validated questionnaire was used to collect data. Results: Participants reported that they are knowledgeable about sperm cryopreservation and gonadotropin-releasing hormone agonists use but not other methods of fertility preservation. About 94% of the participants reported that they need more knowledge about fertility preservation. More than half of the participants had never encountered cancer patients who used ovarian cryopreservation, testicular tissue cryopreservation, in vitro fertilization with embryo cryopreservation and oocyte cryopreservation. The majority (78%) agreed that discussing fertility preservation with newly diagnosed cancer patients is a high priority. Conclusions: Oncologists in Oman are supportive of fertility preservation. The lack of knowledge and unavailability of well-structured fertility preservation services in Oman hinders the initiation of fertility preservation discussions. Exceptional achievement in the early diagnosis of cancer and its treatment causes an increase in life expectancy and a 5-year survival rate. Because of this phenomenal progress, there is a growing population of people who, while being cancer-free for several years, are concerned about the long-term impact of cancer or its treatments on their health, survival, and quality of life . Chemotherapy and radiotherapy are the two most common cancer treatment modalities. Although these treatment modalities are playing a major role in the treatment of cancer, the side effects of chemotherapy and radiotherapy are unpreventable . PrematuGiven the growing number of survivors, it is time to coin the term “fertility preservation”, which refers to the maintenance of fertility in cancer patients after treatment. Fertility preservation options vary between male and female patients as well as between young and adult patients. Male fertility can be preserved by hormonal suppression, testicular sperm extraction, sperm cryopreservation, and testicular tissue preservation. Sperm cryopreservation is by far the simplest and commonest standard technique for post-pubertal males. Testicular tissue freezing and testicular sperm extraction are choices for pre-pubertal boys or men who are sperm deficient. In animal research, germ cell transplantation, germ cell maturation, and stem-cell-to-germ-cell progression have all shown potentials in preserving male fertility . FemalesLoss of infertility has a devastating and long-lasting emotional impact on younger adults ,13,14. FCounseling about cancer, its treatment, and fertility preservation should be given to all cancer patients, especially those who are at their reproductive age. Unfortunately, published works in this regard showed a lack of practice. Fertility preservation was not discussed with patients, and this topic was not included in the medical training programs . FurtherMany cancer survivors would prefer having their own biological child, and they are interested in preserving their fertility ,20,21. EFewer studies are reported to investigate the fertility preservation in developing countries. A lack of awareness among clinicians, patients and caregivers was reported. Moreover, the insufficiency of financial support, absence of service and funding were reported as major barriers to practice fertility preservation in this part of the world. The engagement of stakeholders as well as collaboration and support from developed world countries were recommended . In OmanThis study was conducted at Sultan Qaboos University in Oman from June 2021 to December 2021. This study involved oncologists, treating cancer patients and/or patients receiving chemotherapy and radiotherapy, from four different hospitals and centers.The questionnaire used in this study was adapted from a published article by Adams et al. with minor changes . Three eThe questionnaire is divided into two sections. The first section covers the participants’ demographic information, medical training and practice information. The second section investigated knowledge, current practice, barriers to fertility preservation, perceptions, and attitudes toward FP. The target participants of this study were all the oncologists dealing with cancer patients in four centers and hospitals treating patients with oncological diseases in Oman.Information about the number of oncologists, their specialties, and their contact was obtained with the help of our collaborators in the hospitals and centers included in the study. The cohort of concern  includes 71 oncologists. Our sample size is calculated to be 61 participants with a 95% confidence level and a 5% margin of error. To increase the response rate, all 71 oncologists were invited to participate in the study via an email that included a detailed description of the project’s objectives and goals and contained a link to the questionnaire. Where feasible, face-to-face meetings were used in addition to the email invitations to increase the response rate. The Medical Research Ethics Committee, College of Medicine and Health Sciences, Sultan Qaboos University (REF.NO.SQU-EC/260/2020) approved this study. ®). Descriptive statistics were used to present the demographic data and questionnaire items. In addition, the Chi-Square test was used to compare the categorical variables. For the unequal distribution of answers in some question, some items were turned into dichotomous items. For all statistical tests, a p-value < 0.05 was considered to be statistically significant. The qualitative comments analysis was abandoned, as the information provided did not add to that obtained by the survey.The data analysis was performed using Statistical Package for the Social Sciences software  were older than 40 years old.The majority (65%) of the oncologists that participated in the study were consultants, and about 62% of them reported that they have gone through personal experience of cancer and 94% reported that they have children . p-value = 0.49 and 0.53, respectively).Most of the participants reported that they were knowledgeable mainly about sperm cryopreservation (62%) and pretreatment with GnRH agonists (47%). On the other hand, few participants were aware about ovarian tissue cryopreservation (35.3%) and oocyte cryopreservation (27%) . About 9Participants were asked about the frequency of encountering cancer patients who underwent different methods for FP. They reported that they encountered cancer patients who have used sperm cryopreservation (34%) and pretreatment with GnRH agonist (38%) most often. On the other hand, most physicians reported that they had never encountered a cancer patient who used ovarian tissue cryopreservation (66%), testicular tissue cryopreservation (65%), in vitro fertilization with embryo cryopreservation (59%), and oocyte cryopreservation (59%). About 71% of participants reported that they usually and/or always check with their patients the importance of their future fertility for them. Taking into consideration the desire of the patients for future fertility while planning patient’s treatment regimen is reported by 77% of the participants. Only four participants (12%) reported that they provided their patients with a written information about fertility preservation. The consultation of a fertility specialist or reproductive endocrinologist with questions about the patient’s fertility issues is reported by only four of the participants (12%). Nine of the participants (27%) reported always referring patients for fertility issues discussion with fertility specialists or reproductive endocrinologist prior to or after the cancer treatment. Participants reported that they discuss sperm cryopreservation (77%) and ovarian function suppression with LH-RH agonists (47%) more frequently compared to other methods of fertility preservation. Discussing and providing complete information about FP is one of the patient’s rights. There are many ways to provide cancer patients with enough information regarding the reservation of their fertility. About 68% of the participants believed that referring patients to a fertility specialist would significantly increase their knowledge of their fertility issues. Around 59% of the participants believed that providing educational documents about fertility would be beneficial for the patients to read. In addition, establishing a collaboration between hospitals and fertility centers to disseminate information and knowledge to the patient was reported to be helpful by 53% of the participants. Of the participants, 47% emphasized that continuing education of the patients is of great importance. Moreover, discussing the impact of cancer treatment on patients’ future fertility and providing patients with written information about FP are the participants’ preferred approaches to provide information about FP for those patients.About 65% of the participants reported that they consider 40 as the upper age limit they would consider giving FP counseling and advice to a female patient. However, about 21% of them reported that they would consider providing FP advice to a male patient up to the age of 55. The majority of the participants seemed to be supportive and concerned about FP. The majority of them (78%) agree that discussing fertility preservation with newly diagnosed cancer patients is a high priority. However, 59% reported that treating the primary cancer is more important than FP. About 78% reported that they feel comfortable discussing fertility preservation with their patients. When they were asked about the FP success rate, 35% neither agree nor disagreed that the success rate of fertility preservation options is good enough to make it a viable option for their patients . OncologAbout 62% reported that they think gender is playing a role, and 27% of them thought that women are more concerned. Regarding socioeconomic states, 65% agreed that both high and low socioeconomic states patients are concerned about their future fertility, but 27% reported that the high socioeconomic state is more concerned. Lastly, regarding educational level, participants thought that patients of all educational levels are concerned about their future fertility.The majority of the participants (82%) reported that a lack of infrastructure of fertility services is a major barrier to initiate fertility preservation discussion with their patients. About 59% reported that their limited knowledge about fertility preservation options plays a major role. The priority to treat the cancer, cancer characteristics and prognosis are reported as barriers as well.Patients’ desire to not discuss fertility preservation is reported by 88% of participants as a barrier hindering the initiation of FP discussion. Moreover, patient’s age is also playing a role here, as 74% of the participants reported that treating patients who aged more than 40 years might influence their initiation of FP discussion. Having emotionally distressed patients at the time of the consultation is reported to hinder the initiation of FP discussion by 71% of the participants. About 65% of the participants reported that treating a patient who has a child or children before influences their decision in initiating FP discussion .Chemotherapy and radiotherapy for cancer patients negatively impact ovarian and testicular functions, especially those involving the whole body or mainly the pelvic region . FP has Nearly half, 48%, of the participants responded to the questionnaire, indicating a substantial interest from some oncologists in FP. An important result that emerged from the data of this study is that a vast majority (94%) of our participants indicated that they need more knowledge about FP. This was a very high percentage compared to a study conducted in Japan in which 36% of participants expressed the same . MultiplRegarding the knowledge about different fertility preservation methods, the most knowledgeable fertility preservation methods were sperm cryopreservation 68% and pretreatment with GnRH agonists 47%. Furthermore, this has a complete agreement with previous studies ,25. DespMost of the participants reported that they encountered patients who had used pretreatment with GnRH agonists 38% and sperm cryopreservation 34% most often. Furthermore, this is not surprising, because they reported before that they were knowledgeable mainly in those two methods. Moreover, when recalling the history of FP, the first live birth using sperm cryopreservation was reported in 1953 compared to ovarian tissue cryopreservation, which is a recent milestone in the field . AccordiOur study showed that oncologists seemed supportive of FP, with 78% agreeing or strongly agreeing that FP is a high priority to discuss with their newly diagnosed cancer patients. These findings go hand in hand with what is reported in Adam et al. Moreover, 78% of the oncologists reported that they feel comfortable discussing fertility preservation with their patients, which is a higher percentage than what was reported in Adams et al. (65%). However, 59% also agreed that treating the primary cancer was more important than FP, which is comparable to what had been reported in Adams et al. that 67% of their participants agreed with same statement. Owing this inconsistency to one of the barriers, which is the limited knowledge about fertility preservation options. Given that the majority of participants believed that treating the main cancer is more crucial than preserving fertility, it is hoped that the clinician’s understandable attitude does not impede patients from obtaining the information they need to make an informed decision. For the success rate of fertility preservation methods, 53% agreed or strongly agreed with the statement “success rates of FP are good enough to make it a viable option”. This is comparable to what had been reported in Adam et al. . HoweverIt is imperative to provide accurate and up-to-date information, particularly on a subject as complex as FP, for those patients, which would help them make more informed choices regarding their fertility. Genetic counseling should be included when providing fertility preservation counseling to patients with hereditary cancer syndrome due to their peculiar needs. Personalized counseling that explains the risk of transmission of their genetic cancer susceptibility to their offspring is crucial. There are different ways to provide cancer patients with enough knowledge about FP. Most of our participants preferred referring their patients to a fertility specialist, 68%. When a patient is diagnosed with cancer, and before treatment begins, early referral to a fertility specialist is vital for maximizing the chances of fertility preservation . This waConcerning information provision, more than half (59%) of the participants believe that providing educational documents about fertility for the patients to read would be beneficial. However, many factors may explain why some physicians do not provide their patients with written documents. We assume that the hospitals’ lack of structured written information for their physicians will impact their ability to provide written information. Another aspect is that due to their lack of understanding of FP, physicians will supply any information they can regarding FP to their patients, even if the information is inaccurate or not up to date. According to another survey, about 70% of physicians believe that ensuring prolonged patient survival is more essential than FP . As a reHaving a higher percentage of oncologists who thought that women were more concerned than men about preserving their fertility can be explained by the woman’s ovaries containing a finite number of primordial follicles (termed ovarian reserve) that declines over time and at a higher rate when exposed to chemotherapy. A higher percentage of oncologists thought that patients with high socioeconomic status were more concerned than those with low socioeconomic about preserving their fertility. Few studies investigating the impact of socioeconomic status on FP showed similar results, as FP options are expensive, and high socioeconomic status patients could afford it ,35. RegaAs seen from oncologists’ responses, the existing infrastructure of fertility services seemed to be perceived as a major barrier to providing FP by participants. These findings are in contrast to Adams et al. and Forman et al., who found that fertility services did not seem to be perceived as a major barrier, as the vast majority of participants in their studies reported FP options were available within their trust ,26. ThisThe vast majority of our participants reported that the priority to treat the cancer is influencing them to some extent or a large extent. In contrast, in many published studies, it is found that the discussion of FP has medium to low priority for the oncologists. About half of the oncologists reported their own lack of knowledge about FP as a determining factor. That is agreed with what is reported by Adams et al. . Given the dearth of knowledge reported by the majority of oncologists, it may come as a surprise that the majority claimed they were comfortable addressing FP with patients and that it was a priority. However, addressing the impact of cancer therapy on fertility does not necessitate a thorough understanding of fertility; rather, it is probably more important to evaluate whether FP is appropriate for the woman or couple in question and to know where to refer. Nonetheless, the findings raise the question of what frequent fertility chats might entail and what information might be provided. It would also be fascinating to study how patients interpret this information flow, and additional research is needed to gain a better understanding of their viewpoints. The vast majority stated that cancer prognosis plays a major role. Forman et al. agreed with that, as 88% of their respondents reported that their decision would be influenced by the patients’ poor prognosis . HoweverAbout 88% of the participants stated that having a patient who does not want to discuss fertility preservation is a determining factor for FP discussion. A study published by King et al. participants stated that they would not talk about FP options unless the patients show that they are interested . The vasThis study presents for the first time the awareness level of oncologists in Oman about fertility preservation for patients with cancer. Overall, the study presented an encouraging picture, with the majority of oncologists seeming supportive of FP and feeling comfortable discussing fertility preservation with their patients. However, the study showed that there is a lack of knowledge and a lack of fertility service in Oman, which may be considered major obstacles preventing oncologists from discussing fertility preservation with their patients. The overall lack of knowledge of FP methods suggests that oncologists may benefit from further education and information about current reproductive medicine procedures available.Providing cancer patients with needed information related to their fertility is vital. Therefore, collaboration among different gulf region countries or Arab countries and other parts of the world with advanced care in this regard is needed to formulate a standard guideline for physicians when dealing with cancer patients, including fertility preservation.


Increased oxidative stress and inflammatory response are risk factors for kidney and cardiovascular diseases in patients with hyperuricemia. Uric acid (UA) has been reported to cause inflammation and oxidative damage in cells by inhibiting the nuclear factor E2-related factor 2 (Nrf2) pathway. Notably, Simvastatin (SIM) can regulate the Nrf2 pathway, but whether SIM can regulate inflammatory response and oxidative stress in vascular endothelial cells induced by high UA via this pathway has not been clarified.To demonstrate this speculation, cell activity, as well as apoptosis, was estimated employing CCK-8 and TUNEL, respectively. Indicators of oxidative stress and inflammation were assessed by related kits and western blotting. Subsequently, the effects of SIM on signaling pathways were examined using western blotting.The result showed that after UA exposure, oxidative stress was activated and inflammation was increased, and SIM could reverse this trend. Meanwhile, SIM could inhibit high UA-induced apoptosis. In addition, western blotting results showed that SIM reversed the down-regulation of the expression of Nrf2 pathway-related proteins caused by high UA. SIM alleviated the inflammatory response as well as inhibiting oxidative stress through the Nrf2 pathway, thereby attenuating high UA-induced vascular endothelial cell injury. Hyperuricemia is caused by decreasing or excessive excretion of uric acid (UA) , 2. PrevStatins have been extensively applied in clinical trials due to their numerous effects in addition to lipid-lowering effects . They prHowever, the protective effect of SIM on vascular endothelial cells has not yet been reported. Nuclear factor erythroid 2-related factor 2 (Nrf2), which is a redox-sensitive transcription factor, is the central control factor of NADH Dehydrogenase, Quinone 1 (NQO1) expression under normal homeostasis and oxidative stress , 14. ExtThis study was designed to investigate the possible role of SIM through the Nrf2 signaling pathway in an HUA-induced vascular endothelial cell model.Cell culture2 (16). An in vitro model of hyperuricemia was established by stimulation with 100 μg/ml UA , and subsequently, HUVECs were administrated with 0.2 μM SIM . ML385 , an Nrf2 inhibitor, was used to pretreat the cells for one hour which were then cultured with UA and/or SIM for 24 hr.The human umbilical vein endothelial cells (HUVECs) provided by American Type Culture Collection were plated in endothelial cell medium which was supplemented with 5% FBS, 1% penicillin/streptomycin solution, as well as 1% endothelial cell growth addition  and was placed at 37 °C with 5% COCell vitality assayThe inoculation of HUVECs in 96-well plates was implemented for cell cultivation with UA and/or SIM for 24 hr. Subsequently, 10 μl cell counting kit-8 (CCK-8) solution  was put into every well to further foster the cells. With the aid of an enzyme marker , the absorbance was tested on the premise of λ = 450 nm .Detection of inflammatory cytokinesTo determine the levels of inflammatory cytokines, ELISA was applied. The levels of interleukin-1β (IL-1β), as well as interleukin-18 (IL-18), were examined with an ELISA kit  in light of standard protocol.Measurement of oxidative stress indicatorsThe activity of oxidative stress indicators reactive oxygen species (ROS), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT), as well as malondialdehyde (MDA) in HUVECs was tested employing related kits  in line with standard specifications.Western blotting assayglutathione peroxidase 1 , or anti-GAPDH  was implemented at 4 °C, after which the probe with proper secondary antibody was used. The visualization of protein blots was tracked utilizing the ECL kit , and the analysis was conducted with the application of Image-Pro Plus version 6.0 software  , anti-Bax , anti-Nrf2 , anti-NQO-1 , anti-HO-1 , anti- s, Inc.) .TUNEL assayTerminal Deoxynucleotidyl Transferase mediated dUTP Nick-End Labeling (TUNEL) was adopted for the estimation of apoptotic cell death. Initially, cells were subjected to fixation with 3.7% paraformaldehyde as well as permeabilization with 0.1% Triton X-100. Subsequently, the cultivation of samples with TUNEL assay solution was implemented under light-proof conditions at 37 °C for one hour. Following nuclear staining with DAPI and addition of an anti-fading solution, all samples were photographed at random for observation adopting a fluorescence microscope  .Reverse-transcriptase-polymerase chain reaction (RT-PCR) Total RNA was isolated from the cells with Trizol (Invitrogen USA). Revert Aid (TM) first strand cDNA synthesis kit  was used for RNA reverse transcription. The sequences were as follows: Tumor necrosis factor alpha (TNF-α) sense: 5′-CTGGGCAGGTCTACTTTGGG-3′, antisense: 5′- CTGGAGGCCCCAGTTTGAAT-3′; IL-18 sense: 5′- ATCGCTTCCTCTCGCAACAA-3′, antisense: 5′- GAGGCCGATTTCCTTGGTCA-3′; IL-1β sense: 5′- CCAAACCTCTTCGAGGCACA -3′, antisense: 5′- AGCCATCATTTCACTGGCGA -3′; IL-6 sense: 5′-GTCCAGTTGCCTTCTCCCTGG-3′, antisense: 5′- CCCATGCTACATTTGCCGAAG-3′; GAPDH sense: 5′- AATGGGCAGCCGTTAGGAAA-3′, antisense: 5′- GCGCCCAATACGACCAAATC-3′.Statistical analysisP-values<0.05 meant statistical significance. All experiments were repeated more than 3 times.Data displayed in the form of mean ± SD was analyzed with GraphPad Prism 8.0 software . Comparisons among different groups were demonstrated by one-way ANOVA as well as Tukey’s test. Determination of the application doses of UA and SIMThe application doses of UA and SIM were determined by examination of HUVECs activity. The CCK-8 results showed that the cell viability was 59% and 32% when treated by UA at concentrations of 100 μg/ml and 200 μg/ml, individually, both of which greatly differed from that in the control group . MeanwhiSIM inhibits oxidative stress and inflammation in HUVECs induced by HUAin vitro model of hyperuricemia was established by stimulation with 100 μg/ml UA, and subsequently, HUVECs were administrated with 0.2 μM SIM. The cell activity results showed that co-administration of UA and SIM significantly increased cell activity when compared with that in cells induced by HUA (UA group) (An A group) . At the A group) . The oxiA group) . As depiSIM inhibits apoptosis in HUVECs induced by HUAResults obtained from TUNEL suggested that in contrast with the control group, HUVECs apoptosis in the UA group was prominently exacerbated . HoweverSIM activates the Nrf2 pathway in HUVECs induced by HUAThe effects of UA and SIM on the Nrf2 signaling pathway were then investigated . In contSIM inhibits oxidative stress, inflammation, and apoptosis by regulating the Nrf2 pathwayCCK8 results showed that compared with the UA+SIM group, cell viability was decreased . The resAdditionally, results obtained from TUNEL demonstrated that in contrast with the UA+SIM group, the inhibitory effects of SIM on apoptosis were greatly diminished by inhibition of the Nrf2 pathway by ML385 . And resin vivo (in vivo (Hyperuricemia is produced by high levels of UA in vivo . HUA hasin vivo . Notably(in vivo  has been(in vivo , diabete(in vivo , breast (in vivo -26, as w(in vivo . More im(in vivo . Thus, dIn the present study, the HUA-induced hyperuricemia model was successfully constructed in HUVECs. Subsequently, the effect of SIM on HUA-induced HUVECs was evaluated by the changes in oxidative stress levels and the release of inflammatory cytokines. The results showed that SIM conspicuously repressed apoptosis as well as the levels of IL-1β and IL-18 in HUVECs. Besides, SIM also protected against HUA-elicited oxidative stress, thus mitigating HUA-induced injury in HUVECs.Subsequently, the downstream mechanism of SIM was investigated. Nrf2 acts as a key transcription factor in regulating cellular responses to oxidative stress and inflammation . In anotet al. found that SIM could attenuate TLR4-mediated inflammatory damage and defend against oxidative stress through the Nrf2 signaling pathway in a rat traumatic brain injury model, thus playing a neuroprotective role against traumatic brain injury (Notably, SIM, which is a typical statin with good tolerability, has been increasingly reported . It was n injury . The pren injury . These sSubsequently, the proteins related to Nrf2 signaling pathway were detected. And the results demonstrated that SIM could enhance the contents of Nrf2, NQO-1, as well as HO-1 in HUVECs induced by HUA, which was in line with the previous finding that SIM alleviated oxidative stress-induced renal ischemia/reperfusion injury by targeting the Nrf2/HO-1 pathway . MeanwhiNevertheless, there are also some limitations in this work. Other possible signaling pathways that may be regulated by SIM will be further discussed in future experiments. In addition, other cell lines are needed to further verify our experimental results in addition to HUVECs. In short, the present study further supported the potential value of SIM for the treatment of hyperuricemia.W W and X C conceived and designed the study. X C and L X performed experiments. W W wrote the paper. W W, X C, and L X reviewed and edited the manuscript. W W, X C, and L X confirm the authenticity of all the raw data. All authors have read and approved the final manuscript.SIM reduces high uric acid-induced oxidative stress in vascular endothelial cells. SIM reduces the high uric acid-induced inflammatory response in vascular endothelial cells. SIM activates the Nrf2 signaling pathway.This article does not cover animal experiments and human experiments, thus ethical approval is not required.Personal funds from authors.The authors declare that they have no competing interests.