Histopathological and Immunohistochemical Characterization of Sebaceous Adenoma and Epithelioma in Dogs.

Sebaceous gland tumors are neoplasms originating from the sebaceous gland and are the third most common type of skin tumor, accounting for 21-35% of all cutaneous neoplasms in dogs. According to their histopathological characteristics, sebaceous gland tumors can be classified into adenoma as a benign tumor and epithelioma as a malignant tumor. Sebaceous epithelioma is distinguished from sebaceous adenoma by containing 90% or more reserve cells. However, this simple numerical criterion is insufficient to histologically distinguish between epitheliomas and adenomas. In addition, sebaceoma in humans, a similar tumor to sebaceous epithelioma, is a term used for tumors with more than 50% of reserve cells, unlike epithelioma. Therefore, we aimed to compare and characterize the histological and immunohistochemical profiles of comprehensive sebaceous adenoma, epithelioma, and borderline tumors that have more than 50% but less than 90% of reserve cells. A total of 14 canine sebaceous tumors were diagnosed as seven adenomas, four borderline tumors, and three epitheliomas. Histologically, the sebaceous adenomas showed nodules consisting of mature sebocytes surrounded by monolayer basaloid cells. In contrast, the portion of the reserve cells was increased, the portion of lipidized cells was decreased, and the majority of lipidized cells were found to be immature in sebaceous epithelioma. In the sebaceous adenomas, necrosis was not observed and mitotic figures were rarely seen. However, necrosis and mitotic figures were highly frequent in both borderline tumor and sebaceous epithelioma. Immunohistochemistry revealed that borderline tumor and sebaceous epithelioma showed significantly higher expression against Ki-67 than sebaceous adenoma. We conclude that it is more accurate to employ the cut-off value of 50% reserve cells in humans rather than the current 90% reserve cells for classifying sebaceous gland tumors in dogs, thereby providing new insight into the characterization of the sebaceous gland tumors.

Reappraisal of the morphological and morphometric study of the psoas minor muscle with clinical and developmental insights: cadaveric study.

BACKGROUND: The Psoas Minor (PMi) is the most unstable muscle of the psoas group of the posterior abdominal muscle. This muscle has a fusiform shape and consists of a short fusiform belly continuing distally as a long tendon inserted on the pecten pubis and the iliopectineal arch. The present study was conducted to obtain more detailed information about the muscle and to expand knowledge about its morphology and morphometry. MATERIALS AND METHODS: The posterior abdominal wall of 30 adult cadavers was dissected. Anatomical variabilities in origin, insertion, length, width, and muscle-to-cone ratio were measured when PMi was found. The data collected was interpreted descriptively. RESULTS: PMi was found in 12 cases, ten bilateral and two unilateral. The origin was constant in all cases and, except for three cases, extended into the iliac fascia and the iliopubic eminence. Morphometric analysis revealed that the average length of the proximal muscle belly and distal tendons was 4.52 ± 1.35 cm and 13.05 ± 0.90 cm, respectively. The mean width of the muscle belly was 1.71 ± 0.17 cm, and that of the tendon was 0.47 ± 0.10 cm. On average, the muscle belly occupied the proximal 33.71 ± 6.15% of the total musculotendinous unit. CONCLUSIONS: Findings confirm the inconsistency of PMi in the study population. Morphological variations became more evident as the tendon approached the insertion level. The muscle's distal attachment to the iliac fascia may partially control the position, mechanical stability of the underlying iliopsoas and this circumstantial function may be clinically related to iliopsoas inflammation and pathology. However, further studies recommended to determine biomechanical validity and clinical applicability of this vestigial muscle in human.

The Perspectives of Platelet Proteomics in Health and Disease.

Cardiovascular thromboembolic diseases and cancer continue to be a leading cause of death and disability worldwide. Therefore, it is crucial to advance their diagnoses and treatment in the context of individualized medicine. However, the disease specificity of the currently available markers is limited. Based on analyses of a subset of peptides and matching proteins in disease vs. healthy platelets, scientists have recently shown that focused platelet proteomics enables the quantification of disease-specific biomarkers in humans. In this review, we explored the potential of accurate platelet proteomic research, which is required to identify novel diagnostic and pharmaceutical targets by comprehending the proteome variety of healthy individuals and patients for personalized and precision medicine.

Artemisinin resistance in P. falciparum: probing the interacting partners of Kelch13 protein in parasite.

OBJECTIVES: Artemisinin (ART) resistance in Plasmodium is threatening the artemisinin combination therapies-the first line of defence against malaria. ART resistance has been established to be mediated by the Plasmodium Kelch13 (PfK13) protein. For the crucial role of PfK13 in multiple pathways of the Plasmodium life cycle and ART resistance, it is imperative that we investigate its interacting partners. METHODS: We recombinantly expressed PfK13-p (Bric a brac/Poxvirus and zinc finger and propeller domains), generating anti-PfK13-p antibodies to perform co-immunoprecipitation assays and probed PfK13 interacting partners. Surface plasmon resonance and pull-down assays were performed to establish physical interactions of representative proteins with PfK13-p. RESULTS: The co-immunoprecipitation assays identified 17 proteins with distinct functions in the parasite life cycle- protein folding, cellular metabolism, and protein binding and invasion. In addition to the overlap with previously identified proteins, our study identified 10 unique proteins. Fructose-biphosphate aldolase and heat shock protein 70 demonstrated strong biophysical interaction with PfK13-p, with KD values of 6.6 µM and 7.6 µM, respectively. Additionally, Plasmodium merozoite surface protein 1 formed a complex with PfK13-p, which is evident from the pull-down assay. CONCLUSION: This study adds to our knowledge of the PfK13 protein in mediating ART resistance by identifying new PfK13 interacting partners. Three representative proteins-fructose-biphosphate aldolase, heat shock protein 70, and merozoite surface protein 1-demonstrated clear evidence of biophysical interactions with PfK13-p. However, elucidation of the functional relevance of these physical interactions are crucial in context of PfK13 role in ART resistance.

Shedding Light on the Cell Biology of Platelet-Derived Extracellular Vesicles and Their Biomedical Applications.

EVs are membranous subcellular structures originating from various cells, including platelets which consist of biomolecules that can modify the target cell's pathophysiological functions including inflammation, cell communication, coagulation, and metastasis. EVs, which are known to allow the transmission of a wide range of molecules between cells, are gaining popularity in the fields of subcellular treatment, regenerative medicine, and drug delivery. PEVs are the most abundant EVs in circulation, being produced by platelet activation, and are considered to have a significant role in coagulation. PEV cargo is extremely diverse, containing lipids, proteins, nucleic acids, and organelles depending on the condition that induced their release and can regulate a wide range of biological activities. PEVs, unlike platelets, can overcome tissue barriers, allowing platelet-derived contents to be transferred to target cells and organs that platelets cannot reach. Their isolation, characterization, and therapeutic efficacy, on the other hand, are poorly understood. This review summarizes the technical elements of PEV isolation and characterization methods as well as the pathophysiological role of PEVs, including therapeutic potential and translational possibility in diverse disciplines.

Role of Prednisolone in Platelet Activation by Inhibiting TxA2 Generation through the Regulation of cPLA2 Phosphorylation.

Glucocorticoids have been commonly used in the treatment of inflammation and immune-mediated diseases in human beings and small animals such as cats and dogs. However, excessive use can lead to Cushing's syndrome along with several thrombotic and cardiovascular diseases. Although it is well-known that glucocorticoids exert a significant effect on coagulation, the effect of cortisol on platelet function is much less clear. Thus, we aimed to study the effects of prednisolone, one of the commonly used glucocorticoids, on the regulation of platelet function using murine platelets. We first evaluated the concentration-dependent effect of prednisolone on 2-MeSADP-induced platelet function and found that the 2-MeSADP-induced secondary wave of aggregation and dense granule secretion were completely inhibited from 500 nM prednisolone. Since 2-MeSADP-induced secretion and the resultant secondary wave of aggregation are mediated by TxA2 generation, this result suggested a role of prednisolone in platelet TxA2 generation. Consistently, prednisolone did not affect the 2-MeSADP-induced aggregation in aspirinated platelets, where the secondary wave of aggregation and secretion were blocked by eliminating the contribution of TxA2 generation by aspirin. In addition, thrombin-induced platelet aggregation and secretion were inhibited in the presence of prednisolone by inhibiting the positive-feedback effect of TxA2 generation on platelet function. Furthermore, prednisolone completely inhibited 2-MeSADP-induced TxA2 generation, confirming the role of prednisolone in TxA2 generation. Finally, Western blot analysis revealed that prednisolone significantly inhibited 2-MeSADP-induced cytosolic phospholipase A2 (cPLA2) and ERK phosphorylation in non-aspirinated platelets, while only cPLA2 phosphorylation, but not ERK phosphorylation, was significantly inhibited by prednisolone in aspirinated platelets. In conclusion, prednisolone affects platelet function by the inhibition of TxA2 generation through the regulation of cPLA2 phosphorylation, thereby shedding light on its clinical characterization and treatment efficacy in dogs with hypercortisolism in the future.

Antiplatelet Effect of Daphnetin Is Regulated by cPLA2-Mediated Thromboxane A2 Generation in Mice.

Coumarin derivatives have been recognized for their antithrombotic, anti-inflammatory, and antioxidant properties, and daphnetin is one of the natural coumarin derivatives isolated from Daphne Koreana Nakai. Although the pharmacological value of daphnetin is well documented in diverse biological activities, its antithrombotic effect has not been studied to date. Here, we characterized the role and underlying mechanism of daphnetin in the regulation of platelet activation using murine platelets. In order to check the effect of daphnetin on platelet function, we first measured the effect of daphnetin on platelet aggregation and secretion. Collagen-induced platelet aggregation and dense granule secretion were partially inhibited by daphnetin. Interestingly, 2-MeSADP-induced secondary waves of aggregation and secretion were completely inhibited by daphnetin. It is known that 2-MeSADP-induced secretion and the resultant secondary wave of aggregation are mediated by the positive feedback effect of thromboxane A2 (TxA2) generation, suggesting the important role of daphnetin on TxA2 generation in platelets. Consistently, daphnetin did not affect the 2-MeSADP-induced platelet aggregation in aspirinated platelets where the contribution of TxA2 generation was blocked. Additionally, platelet aggregation and secretion induced by a low concentration of thrombin, which is affected by the positive feedback effect of TxA2 generation, were partially inhibited in the presence of daphnetin. Importantly, 2-MeSADP- and thrombin-induced TxA2 generation was significantly inhibited in the presence of daphnetin, confirming the role of daphnetin on TxA2 generation. Finally, daphnetin significantly inhibited 2-MeSADP-induced cytosolic phospholipase A2 (cPLA2) and ERK phosphorylation in non-aspirinated platelets. Only cPLA2 phosphorylation, not ERK phosphorylation, was significantly inhibited by daphnetin in aspirinated platelets. In conclusion, daphnetin plays a critical role in platelet function by inhibiting TxA2 generation through the regulation of cPLA2 phosphorylation.

Fatal Clostridium sporogenes Soft Tissue Polymicrobial Infections in Two Immunocompetent Cases: Case Report.

BACKGROUND: Clostridium sporogenes is reported rarely in literature. Reports from the skin and soft tissue infections are even less, more so in immunocompetent patients. CASE PRESENTATION: Two skin and soft tissue infections with C. sporogenes in immunocompetent patients have been presented in this study. One of the cases was following an electrical burn wound, and the other was following a bedsore. Both patients expired despite antibacterial treatment and debridement. DISCUSSION AND CONCLUSION: C. sporogenes had usually been reported after trauma particularly after penetrating and deep wound infection. More attention should be given to these patients so that the infection can be treated and diagnosed early in suspected anaerobic infections like Clostridium species.

Hand hygiene with interventions: an observational study from a tertiary care institute over 2 years.

Background: Appropriate adherence to hand hygiene (HH) practices by health care workers (HCWs) reduces the transmission of pathogens and subsequently the incidence of hospital acquired infections (HAIs), in health care settings. Strict monitoring and auditing of this simple and cost-effective intervention is very important, as it significantly contributes in reducing the HAIs. Material and methods: A retrospective observational study, evaluating the HH audits from June 2021 till May 2023 in a tertiary health care facility in North India. HH audits were conducted in the ICUs and wards daily, by the trained infection control nurses (ICNs), using direct observation method based on World health organization (WHO) hand hygiene observational forms. HH total adherence (HHTAR), partial adherence (HHPAR) and complete adherence rate (HHCAR) were analyzed in Microsoft Excel sheet. HHTAR rates were compared among different profession, moments and the month wise trend was also observed over the period. Results: A total of 24,740 HH opportunities were observed. The compliance rate for HHCAR, HHPAR and HHTAR were 20.3%, 41.5% and 61.4% respectively. Overall better compliance was reported from the ICUs, profession-specific compliance was highest among nurses (62.8%) and doctors (61.5%). Significant increase in adherence rate was appreciated post intervention 46.1% to 67.3%, (p value < 0.01). Conclusions: Continuous monitoring and reinforcement with timely feedback for intervention and regular auditing is a necessity to improve and maintain the appropriate HH practices among the HCWs. Low- and middle-income countries need to focus more on this simple and promising measure to combat the increasing HAI rates.

Hearing Loss among Children Visiting Department of Otolaryngology and HNS of a Tertiary Care Centre.

Introduction: Hearing loss is defined as the partial or total reduction in auditory acuity. Hearing loss can cause detrimental effects on speech, language, developmental, educational, and cognitive outcomes in children. This study aimed to find out the prevalence of hearing loss among children visiting the Department of Otolaryngology and HNS of a tertiary care centre. Methods: A descriptive cross-sectional study was conducted among children visiting the Department of Otolaryngology and HNS between 1 January 2022 and 31 December 2022 after obtaining ethical approval. All the patients who underwent pure tone evaluation were included in the study. A convenience sampling technique was used. The point estimate was calculated at a 95% Confidence Interval. Results: Among 3051 children, 328 (10.75%) (9.65-11.85, 95% Confidence Interval) had hearing loss. Among children with hearing loss, 170 (51.83%) of children were female. The mean age of children with hearing loss was 13.31±3.39 years. The mean pure tone average among 452 ears with hearing loss was 44.60±17.71 dB. The commonest degree of hearing loss was mild hearing loss 266 (58.85%), and the commonest type was conductive hearing loss 310 (68.58%). Among children with hearing loss, 124 (37.80%) had bilateral hearing loss. Conclusions: The prevalence of hearing loss among children visiting the Department of Otolaryngology and HNS was found to be lower than similar studies done in similar settings. Keywords: audiology; audiometry; hearing loss; outpatients; prevalence.

Chronic Otitis Media among Patients Visiting Community-Based Static Outreach Clinics.

Introduction: Chronic otitis media is a chronic inflammation of the middle ear and mastoid cavity, with recurrent ear discharges or otorrhoea through a tympanic perforation for the past 3 months. It is a common cause of hearing impairment, disability, and poor scholastic performance and can lead to fatal intracranial infections and acute mastoiditis. This study aimed to find out the prevalence of chronic otitis media among patients visiting community-based static outreach clinics. Methods: A descriptive cross-sectional study was conducted among patients visiting the community-based static outreach clinics from 1 January 2017 to 31 December 2019. The ethical approval was taken from the Ethical Review Board. The diagnosis of chronic otitis media was done using otoscopy. The records of patients coming to outreach clinics visiting for ear, nose and throat care were reviewed using a pre-designed study proforma. A systematic random sampling method was used. The point estimate was calculated at a 95% Confidence Interval. Results: Among 385 patients, 37 (9.61%) (6.67-12.55, 95% Confidence Interval) had chronic otitis media. The mean age of patients with chronic otitis media was 27.59±13.24 years, with 28 (75.67%) patients aged between 18-60 years. Among them, 30 (81.08%) had unilateral and 34 (91.89%) had a mucosal type. Conclusions: The prevalence of chronic otitis media was lower than in other studies done in similar settings. Keywords: community health services; otitis media; patients; prevalence.

Effect of Multimodal Intervention (computer based cognitive training, diet and exercise) in comparison to health awareness among older adults with Subjective Cognitive Impairment (MISCI-Trial)-A Pilot Randomized Control Trial.

INTRODUCTION: This study was aimed to investigate the effect of multimodal intervention on the cognitive functions of older adults with subjective cognitive impairment (SCI). MATERIALS AND METHODS: Sixty subjects were randomized 1:1:1:1 to receive either computer based cognitive therapy (CBCT) or CBCT+Mediterranean equivalent diet (MED) or CBCT+MED+ Exercise regime and the control group. The intervention group received supervised CBCT twice a week to have 40 sessions, each of 40 minutes duration, and/ or supervised aerobic and resistive exercise twice a week for 24 weeks and or MED at home under the supervision of a dietician. The control group was provided with health awareness instructions for brain stimulating activities such as sudoku, mental maths, and learning music and new skills. RESULTS: Cognitive functions which was the primary outcome measure were assessed using the Post Graduate Institute Memory Scale (PGI-MS), and Stroop Colour and Word Test at baseline and after 6 months intervention period. As assessed by the PGI-MS, there was significant improvement in domains such as mental balance, attention and concentration, delayed recall, immediate recall, verbal retention of dissimilar pairs, Visual retention, and total score both in the unimodal and multimodal intervention groups. However, the improvement was observed to be the highest in the multimodal intervention group as compared to unimodal group. All the participants completed the trial. CONCLUSION: This pilot randomized control trial indicated that multimodal intervention could be an effective non-pharmacological intervention in individuals with SCI for improving their cognitive functions.

An understanding of coronavirus and exploring the molecular dynamics simulations to find promising candidates against the Mpro of nCoV to combat the COVID-19: A systematic review.

The first infection case of new coronavirus was reported at the end of 2019 and after then, the cases are reported in all nations across the world in a very short period. Further, the regular news of mutations in the virus has made life restricted with appropriate behavior. To date, a new strain (Omicron and its new subvariant Omicron XE) has brought fear amongst us due to a higher trajectory of increase in the number of cases. The researchers thus started giving attention to this viral infection and discovering drug-like candidates to cure the infections. Finding a drug for any viral infection is not an easy task and takes plenty of time. Therefore, computational chemistry/bioinformatics is followed to get promising molecules against viral infection. Molecular dynamics (MD) simulations are being explored to get drug candidates in a short period. The molecules are screened via molecular docking, which provides preliminary information which can be further verified by molecular dynamics (MD) simulations. To understand the change in structure, MD simulations generated several trajectories such as root mean square deviation (RMSD), root mean square fluctuation (RMSF), hydrogen bonding, and radius of gyration for the main protease (Mpro) of the new coronavirus (nCoV) in the presence of small molecules. Additionally, change in free energy for the formation of complex of Mpro of nCoV with the small molecule can be determined by applying molecular mechanics with generalized born and surface area solvation (MM-GBSA). Thus, the promising molecules can be further explored for clinical trials to combat coronavirus disease-19 (COVID-19).

A complete clinico-epidemiological and microbiological profile of candidemia cases in a tertiary-care hospital in Western India.

Objective: To identify different Candida spp along with antifungal susceptibility pattern and risk factors associated with candidemia. Design setting and patients: This retrospective observational study was conducted in a tertiary-care academic hospital in Jaipur, Western India, for 3 years (July 2017-June 2020). Methods: Blood cultures were performed according to standard microbiological methods, and only 1 isolate per patient was included in the study. Isolates of Candida spp were identified using a VITEK-2 automated system and matrix-assisted laser desorption ionization-time of flight mass spectrometry. Antifungal susceptibility tests were performed using the broth microdilution assay according to the Clinical and Laboratory Standards Institute guidelines. Results: Of 3,443 blood cultures received from suspected sepsis cases, candidemia was identified in 95 (2.8%). In addition to Candida tropicalis (n = 36; 38%) and Candida parapsilosis (n = 17; 18%), 10 isolates of Candida auris comprised the fourth most common cause of candidemia. Presence of central venous catheter and diabetes were statistically significant risk factors for development of candidemia by NAC. Resistance to fluconazole was 36%, resistance to voriconazole was 20%, resistance to 5-flucytosine was 4%, and resistance to amphotericin-B was 7%. C. auris isolates were more resistant than other NAC spp. We detected no resistance among the echinocandins. Conclusions: The emergence of highly resistant isolates like C. auris emphasizes the need for constant monitoring of candidemia cases for species identification and routine antifungal susceptibility so that appropriate measures can be taken to reduce the related morbidity and mortality.

An Insight into Recent Advances on Platelet Function in Health and Disease.

Platelets play a variety of roles in vascular biology and are best recognized as primary hemostasis and thrombosis mediators. Platelets have a large number of receptors and secretory molecules that are required for platelet functionality. Upon activation, platelets release multiple substances that have the ability to influence both physiological and pathophysiological processes including inflammation, tissue regeneration and repair, cancer progression, and spreading. The involvement of platelets in the progression and seriousness of a variety of disorders other than thrombosis is still being discovered, especially in the areas of inflammation and the immunological response. This review represents an integrated summary of recent advances on the function of platelets in pathophysiology that connects hemostasis, inflammation, and immunological response in health and disease and suggests that antiplatelet treatment might be used for more than only thrombosis.

The Shape of Nanostructures Encodes Immunomodulation of Carbohydrate Antigen and Vaccine Development.

Gold nanoparticles (AuNPs) have shown remarkable potential for vaccine development, but the influence of the size and shape of nanoparticles modulating the T-cell-dependent carbohydrate antigen processing and immunomodulation is poorly investigated. Here, we described how different shapes and sizes of gold nanostructures carrying adjuvant modulate carbohydrate-based antigen processing in murine dendritic cells (mDCs) and subsequent T-cell activation produce a robust antibody response. As a prototype, CpG-adjuvant-coated spherical and rod- and star-shaped AuNPs were conjugated to the tripodal Tn-glycopeptide antigen to study their DC uptake and activation of T-cells in a DCs/T-cell co-culture assay. Our results showed that the spherical and star-shaped AuNPs displayed relatively weak receptor-mediated uptake and endosomal sequestration; however, they induced a high level of T helper-1 (Th1) biasing immune responses compared with rod-shaped AuNPs. Furthermore, the in vivo administration of AuNPs showed that the small spherical and star-shaped AuNPs induced an effective anti-Tn-glycopeptide immunoglobulin (IgG) antibody response compared with rod-shaped AuNPs. These results indicated that one could obtain superior carbohydrate vaccines by varying the shape and size parameters of nanostructures.

An Insight into GPCR and G-Proteins as Cancer Drivers.

G-protein-coupled receptors (GPCRs) are the largest family of cell surface signaling receptors known to play a crucial role in various physiological functions, including tumor growth and metastasis. Various molecules such as hormones, lipids, peptides, and neurotransmitters activate GPCRs that enable the coupling of these receptors to highly specialized transducer proteins, called G-proteins, and initiate multiple signaling pathways. Integration of these intricate networks of signaling cascades leads to numerous biochemical responses involved in diverse pathophysiological activities, including cancer development. While several studies indicate the role of GPCRs in controlling various aspects of cancer progression such as tumor growth, invasion, migration, survival, and metastasis through its aberrant overexpression, mutations, or increased release of agonists, the explicit mechanisms of the involvement of GPCRs in cancer progression is still puzzling. This review provides an insight into the various responses mediated by GPCRs in the development of cancers, the molecular mechanisms involved and the novel pharmacological approaches currently preferred for the treatment of cancer. Thus, these findings extend the knowledge of GPCRs in cancer cells and help in the identification of therapeutics for cancer patients.

A Cell-Culture Technique to Encode Glyco-Nanoparticles Selectivity.

Nanoparticles (NPs) embedded with bioactive ligands such as carbohydrates, peptides, and nucleic acid have emerged as a potential tool to target biological processes. Traditional in vitro assays performed under statistic conditions may result in non-specific outcome sometimes, mainly because of the sedimentation and self-assembly nature of NPs. Inverted cell-culture assay allows for flexible and accurate detection of the receptor-mediated uptake and cytotoxicity of NPs. By combining this technique with glyco-gold nanoparticles, cellular internalization and cytotoxicity were investigated. Regioselective glycosylation patterns and shapes of the NPs could tune the receptors' binding affinity, resulting in precise cellular uptake of gold nanoparticles (AuNPs). Two cell lines HepG2 and HeLa were probed with galactosamine-embedded fluorescent AuNPs, revealing significant differences in cytotoxicity and uptake mechanism in upright and invert in vitro cell-culture assay, high-specificity toward uptake, and allowing for a rapid screening and optimization technique.

The Predominant Role of Arrestin3 in General GPCR Desensitization in Platelets.

Arrestins in concert with GPCR kinases (GRKs) function in G protein-coupled receptor (GPCR) desensitization in various cells. Therefore, we characterized the functional differences of arrestin3 versus arrestin2 in the regulation of GPCR signaling and its desensitization in platelets using mice lacking arrestin3 and arrestin2. In contrast to arrestin2, platelet aggregation and dense granule secretion induced by 2-MeSADP, U46619, thrombin, and AYPGKF were significantly potentiated in arrestin3-deficient platelets compared to wild-type (WT) platelets, while non-GPCR agonist CRP-induced platelet aggregation and secretion were not affected. Surprisingly, in contrast to GRK6, platelet aggregation induced by the co-stimulation of serotonin and epinephrine was significantly potentiated in arrestin3-deficient platelets, suggesting the central role of arrestin3 in general GPCR desensitization in platelets. In addition, the second challenge of ADP and AYPGKF restored platelet aggregation in arrestin3-deficient platelets but failed to do so in WT and arrestin2-deficient platelets, confirming that arrestin3 contributes to GPCR desensitization. Furthermore, ADP- and AYPGKF-induced Akt and ERK phosphorylation were significantly increased in arrestin3-deficient platelets. Finally, we found that arrestin3 is critical for thrombus formation in vivo. In conclusion, arrestin3, not arrestin2, plays a central role in the regulation of platelet functional responses and thrombus formation through general GPCR desensitization in platelets.

Rational designing of glyco-nanovehicles to target cellular heterogeneity.

The aberrant expression of endocytic epidermal growth factor receptors (EGFRs) in cancer cells has emerged as a key target for therapeutic intervention. Here, we describe for the first time a state-of-the-art design for a heparan sulfate (HS) oligosaccharide-based nanovehicle to target EGFR-overexpressed cancer cells in cellular heterogeneity. An ELISA plate IC50 inhibition assay and surface plasma resonance (SPR) binding assay of structurally well-defined HS oligosaccharides showed that 6-O-sulfation (6-O-S) and 6-O-phosphorylation (6-O-P) of HS tetrasaccharides significantly enhanced EGFR cognate growth factor binding. The conjugation of these HS ligands to multivalent fluorescent gold nanoparticles (AuNPs) enabled the specific and efficient targeting of EGFR-overexpressed cancer cells. In addition, this heparinoid-nanovehicle exhibited selective homing to NPs in cancer cells in three-dimensional (3D) coculture spheroids, thus providing a novel target for cancer therapy and diagnostics in the tumor microenvironment (TME).