Objective:To explore the clinical and pathological features and survival outcomes of patients with early-onset intrahepatic cholangiocarcinoma (EOICC).Methods:This is a multicenter, retrospective cohort study. Data of 1 160 intrahepatic cholangiocarcinoma patients undergoing radical resection in 14 tertiary Grade A hospitals in China from January 2010 to November 2021 were retrospectively collected. The cohort included 632 males and 528 females, aged( M (IQR)) 61 (14) years (range: 22 to 93 years). ICC aged ≤50 years at the time of diagnosis was defined as EOICC and >50 years as late-onset intrahepatic cholangiocarcinoma (LOICC). Of these, there were 247 cases in the EOICC group and 913 cases in the LOICC. The clinical and pathological characteristics of both groups were analyzed and compared using the independent sample t-test, Mann-Whitney U test or Kaplan-Meier method. Univariate and multivariate Cox regression models for patient outcomes were constructed and forest graphed. Results:Compared with the patients in the LOICC group, patients in the EOICC group had lower carcinoembryonic antigen levels (2.5(4.0) μg/L vs. 3.1(5.2)μg/L, U=124 899, P=0.009) and CA19-9 level (63.4(524.7)U/ml vs. 77.9(611.3)U/ml, U=120 320, P=0.013), higher levels of ALT (29(35)U/L vs. 24(26)U/L, U=101 214, P=0.013), a lower score of the Eastern US Cooperative Oncology Group (0 score patients: 54.7% vs. 44.1%, χ2=12.472, P=0.014), higher TNM stage ( χ2=11.807, P=0.038), and proportion of lymph node dissection (62.3% vs. 54.1%, χ2=5.355, P=0.021). Patients in the two groups in sex, first diagnosis symptoms, intrahepatic bile duct stone history, nail protein, albumin, total bilirubin, transaminase, liver function Child-Pugh grade, T stage, stage, N stage, preoperative laparoscopic exploration proportion, tumor diameter, vascular invasion proportion, differentiation, margin, intraoperative bleeding, postoperative complications, postoperative hospital days were no statistical significance (all P>0.05). Patients in the EOICC group had better outcomes than the LOICC group (median survival time: 29.7 months vs. 25.0 months, 3-year overall survival: 45.1% vs. 37.8%, P=0.027). Conclusion:EOICC patients are better than LOICC patients in carcinoembryonic antigen, CA19-9, ALT, physical strength status and TNM stage, and the long-term prognosis is also better than LOICC patients.

Antibody humanization is critical to reduce immunogenicity and enhance efficacy in the preclinical phase of the development of therapeutic antibodies originated from animal models.Computational suggestions have long been desired,but available tools focused on immunogenicity calculation of whole antibody sequences and sequence segments,missing the individual residue sites.This study introduces Site-specific Immunogenicity for Therapeutic Antibody(SITA),a novel computational framework that predicts B-cell immunogenicity score for not only the overall antibody,but also individual residues,based on a comprehensive set of amino acid descriptors characterizing physicochemical and spatial features for antibody structures.A transfer-learning-inspired framework was purposely adopted to overcome the scarcity of Antibody-Antibody structural complexes.On an independent testing dataset derived from 13 Antibody-Antibody structural complexes,SITA successfully predicted the epitope sites for Antibody-Antibody structures with a receiver operating characteristic(ROC)-area unver the ROC curve(AUC)of 0.85 and a precision-recall(PR)-AUC of 0.305 at the residue level.Furthermore,the SITA score can significantly distinguish immunogenicity levels of whole human antibodies,therapeutic antibodies and non-human-derived antibodies.More importantly,analysis of an additional 25 thera-peutic antibodies revealed that over 70％of them were detected with decreased immunogenicity after modification compared to their parent variants.Among these,nearly 66％antibodies successfully iden-tified actual modification sites from the top five sites with the highest SITA scores,suggesting the ability of SITA scores for guide the humanization of antibody.Overall,these findings highlight the potential of SITA in optimizing immunogenicity assessments during the process of therapeutic antibody design.

Antibody (Ab) humanization is critical to reduce immunogenicity and enhance efficacy in the preclinical phase of the development of therapeutic Abs originated from animal models. Computational suggestions have long been desired, but available tools focused on immunogenicity calculation of whole Ab sequences and sequence segments, missing the individual residue sites. This study introduces Site-specific Immunogenicity for Therapeutic Antibody (SITA), a novel computational framework that predicts B-cell immunogenicity score for not only the overall antibody, but also individual residues, based on a comprehensive set of amino acid descriptors characterizing physicochemical and spatial features for antibody structures. A transfer-learning-inspired framework was purposely adopted to overcome the scarcity of Ab-Ab structural complexes. On an independent testing dataset derived from 13 Ab-Ab structural complexes, SITA successfully predicted the epitope sites for Ab-Ab structures with a receiver operating characteristic (ROC)-area unver the ROC curve (AUC) of 0.85 and a precision-recall (PR)-AUC of 0.305 at the residue level. Furthermore, the SITA score can significantly distinguish immunogenicity levels of whole human Abs, therapeutic Abs and non-human-derived Abs. More importantly, analysis of an additional 25 therapeutic Abs revealed that over 70% of them were detected with decreased immunogenicity after modification compared to their parent variants. Among these, nearly 66% Abs successfully identified actual modification sites from the top five sites with the highest SITA scores, suggesting the ability of SITA scores for guide the humanization of antibody. Overall, these findings highlight the potential of SITA in optimizing immunogenicity assessments during the process of therapeutic antibody design.

Objective To examine the role and mechanism of clopidogrel in the development of salt-sensitive hypertension.Methods 8-week-old Dahl salt-sensitive(Dahl SS)rats and control salt-resistant(SS13BN)rats were divided randomly into six groups and fed for 8 weeks with normal salt(0.4％NaCl,NS),high salt(8％NaCl,HS),or high salt combined with clopidogrel gavage(8％NaCl+10 mg/(kg·d))clopidogrel,HS+CLO).Arterial systolic blood pressure was measured continuously over 8 weeks by the tail-cuff method,and systolic blood pressure was measured by carotid cannulation after 8 weeks(56 days).Renal histopathology was observed by hematoxylin and eosin staining,and renal inflammatory cell infiltration was detected by immunohistochemistry.Peripheral blood platelet activation and platelet-leukocyte aggregation were analyzed by flow cytometry,and the renal inflammation-related proteins tumor necrosis factor-α,interleukin(IL)-1β,IL-6,and key proteins in the p38MAPK/nuclear factor(NF)-κB signaling pathway were detected by Western blot.Results Compared with the NS group,Dahl SS HS rats had significantly increased blood pressure(P＜0.05),aggravated renal tissue damage,increased inflammatory cell infiltration,increased expression of inflammatory cytokines(P＜0.05),elevated peripheral blood platelet activation(P＜0.05)and platelet-leukocyte aggregation(P＜0.05),and increased expression of p38MAPK/NF-κB signaling pathway proteins(P＜0.05).Clopidogrel effectively alleviated these phenotypes induced by high salt in Dahl SS rats.Conclusions Clopidogrel alleviated high-salt-induced salt-sensitive hypertension and decreased renal inflammatory responses and dysfunction in Dahl SS rats by inhibiting platelet activation and the p38MAPK/NF-κB signaling pathway.

Objective:To investigate the role of umbilical cord mesenchymal stem cell (MSC) -derived nanovesicles in hair regeneration.Methods:(1) Nanovesicles were prepared by continuously extruding umbilical cord MSCs through polycarbonate membranes, and were identified using transmission electron microscopy and nanoparticle tracking analysis. (2) Six C57BL/6 female mice with full-thickness skin wounds were randomly divided into a nanovesicle group (subcutaneously injected with nanovesicles once at the wound margin) and a control group (subcutaneously injected with an equal volume of phosphate-buffered saline [PBS] at the wound margin) ; skin samples were collected on day 16 for hematoxylin-eosin (HE) staining to assess wound healing and hair follicle regeneration. (3) Human hair follicle dermal papilla cells (DPCs) were isolated using a two-step enzyme method; the uptake of PKH26-pre-labeled nanovesicles by DPCs was observed by fluorescence microscopy; the proliferative activity of DPCs co-cultured with nanovesicles was evaluated using cell counting kit-8 (CCK8) and 5-ethynyl-2'-deoxyuridine (EdU) assays. (4) Six healthy C57BL/6 female mice were randomly divided into two groups after anesthesia, and subcutaneously injected with either fluorescent dye DIR-pre-labeled nanovesicles or PBS; an in vivo imaging system was used to observe the uptake and metabolism of nanovesicles in the mouse skin. (5) Twenty-four C57BL/6 female mice with depilated backs were randomly divided into a nanovesicle group (subcutaneously injected with nanovesicles on days 0, 8, and 15) and a control group (subcutaneously injected with an equal volume of PBS at the same time points) ; skin samples were collected on days 4, 18, and 21 for HE staining to analyze differences in hair follicle cycling; transcriptome sequencing was performed on skin samples collected on day 4. Statistical analyses were conducted using the t test. Results:(1) Transmission electron microscopy showed that nanovesicles exhibited a spherical membranous structure with diameters of 141.3 ± 60.0 nm. (2) In 6 C57BL/6 female mice with full-thickness skin wounds, the wound area on day 12 was significantly smaller in the nanovesicle group (1.27 ± 0.50 mm 2) than in the control group (4.13 ± 1.03 mm 2, t = 4.34, P = 0.012). (3) Fluorescence microscopy revealed that nanovesicles were taken up by DPCs within 20 hours; the absorbance of DPCs was significantly higher in the nanovesicle group than in the control group ( t = 20.23, P < 0.001), and the percentage of EdU-positive cells was also significantly higher in the nanovesicle group (49.62% ± 6.45%) than in the control group (37.58% ± 3.42%, t = 3.69, P = 0.006). (4) In vivo imaging of the 6 C57BL/6 female mice showed strong fluorescence in the back of mice in the nanovesicle group on day 0, which markedly decreased by day 8, while no fluorescence was observed in the control group throughout the experiment. (5) Hair follicle cycle experiments on the 24 C57BL/6 female mice with depilated backs showed that the hair follicle length on day 4 after depilation was significantly longer in the nanovesicle group (368.00 ± 63.17 μm) than in the control group (266.90 ± 34.41 μm, t = 9.87, P < 0.001), and the hair bulb diameter was also significantly longer in the nanovesicle group (54.83 ± 10.32 μm) than in the control group (39.12 ± 7.54 μm, t = 16.02, P < 0.001) ; on day 18, the nanovesicle group showed a significantly higher hair follicle density (19.12 ± 0.90) compared with the control group (11.07 ± 1.51, t = 7.92, P = 0.001) ; on day 21, 46.13% ± 8.64% of hair follicles in the nanovesicle group remained in the anagen phase Ⅵ to the catagen phase Ⅱ, and 46.24% ± 3.29% were in the catagen phases Ⅲ to Ⅳ, while 78.89% ± 18.36% of hair follicles in the control group were in the telogen phases Ⅶ to Ⅷ. Transcriptome sequencing showed that differentially expressed genes in the nanovesicle group were significantly positively enriched in the keratinization process (NES = 2.23, P < 0.001) . Conclusion:Umbilical cord MSC-derived nanovesicles could promote the proliferation of DPCs, advance the entry of hair follicles into the anagen phase, delay their entry into the catagen phase, and induce hair regeneration.

Objective:To investigate the effect of verbascoside(VERB)on high-fat diet-induced atherosclerosis(AS)in ApoE-/-mice and the effect on high mobility histone 1(HMGB1)/receptor for glycation end products(RAGE)/nuclear factor κB(NF-κB)pathway.Methods:A total of 90 ApoE-/-mice were randomly divided into normal group,AS group,VERB group,simvastatin group and VERB+pathway activator HMGB1 group,with 18 mice per group.After 8 weeks of group administration,blood and aorta samples were taken.Fasting serum triacylglycerol(TG),total cholesterol(TC)and low density lipoprotein(LDL)levels were deter-mined by automatic biochemical analyzer.Oil red O,HE and TUNEL staining were performed to observe apoptosis of aortic plaque and endothelial cell(EC).Flow cytometry was performed to analyze circulating EC numbers.Immunohistochemistry was performed to analyze the infiltration area of macrophages(CD68+)and T lymphocytes(CD3+)in aortic plaques.Western blot was performed to detect expressions of HMGB1/RAGE/NF-κB pathway,inflammation and adhesion molecules.Results:Compared with normal group,AS group had lipid plaques in arterial intima,thickness of the media was uneven,TG,TC,LDL levels,lession proportion,plaque area,circulating EC number,arterial EC apoptosis rate,macrophage(CD68+)and T lymphocyte(CD3+)infiltration areas,TNF-α,monocyte chemoattractant protein-1(MCP-1),vascular cell adhesion molecule-1(VCAM-1),intercellular adhesion molecule-1(ICAM-1),cytoplasm HMGB1,total HMGB1,total RAGE protein levels and nuclear/total p65 NF-κB levels were increased(P<0.05),while nuclear HMGB1 protein,nuclear/total HMGB1 and cytosolic p65 NF-κB levels were decreased(P<0.05).After VERB or simvastatin intervention,arterial lesions were alleviated,TG,TC,LDL levels,lession proportion,plaque area,circulating EC number,arterial EC apoptosis rate,macrophage(CD68+)and T lymphocyte(CD3+)infiltration areas,TNF-α,MCP-1,VCAM-1,ICAM-1,cytoplasm HMGB1,total HMGB1,RAGE protein levels and nuclear/total p65 NF-κB level were decreased(P<0.05),while nuclear HMGB1 protein,nuclear/total HMGB1 and cytosolic p65 NF-κB levels were increased(P<0.05),and HMGB1 was able to antagonize the protective effect of VERB on AS mice.Conclusion:VERB can inhibit expressions of inflammatory and adhesion fac-tors in arterial plaques in ApoE-/-mice,reduce EC shedding and apoptosis,therefore improve AS symptoms in ApoE-/-mice,and the mechanism may be related to the inhibition of HMGB1/RAGE and NF-κB pathway.

Objective:To investigate the role of umbilical cord mesenchymal stem cell (MSC) -derived nanovesicles in hair regeneration.Methods:(1) Nanovesicles were prepared by continuously extruding umbilical cord MSCs through polycarbonate membranes, and were identified using transmission electron microscopy and nanoparticle tracking analysis. (2) Six C57BL/6 female mice with full-thickness skin wounds were randomly divided into a nanovesicle group (subcutaneously injected with nanovesicles once at the wound margin) and a control group (subcutaneously injected with an equal volume of phosphate-buffered saline [PBS] at the wound margin) ; skin samples were collected on day 16 for hematoxylin-eosin (HE) staining to assess wound healing and hair follicle regeneration. (3) Human hair follicle dermal papilla cells (DPCs) were isolated using a two-step enzyme method; the uptake of PKH26-pre-labeled nanovesicles by DPCs was observed by fluorescence microscopy; the proliferative activity of DPCs co-cultured with nanovesicles was evaluated using cell counting kit-8 (CCK8) and 5-ethynyl-2'-deoxyuridine (EdU) assays. (4) Six healthy C57BL/6 female mice were randomly divided into two groups after anesthesia, and subcutaneously injected with either fluorescent dye DIR-pre-labeled nanovesicles or PBS; an in vivo imaging system was used to observe the uptake and metabolism of nanovesicles in the mouse skin. (5) Twenty-four C57BL/6 female mice with depilated backs were randomly divided into a nanovesicle group (subcutaneously injected with nanovesicles on days 0, 8, and 15) and a control group (subcutaneously injected with an equal volume of PBS at the same time points) ; skin samples were collected on days 4, 18, and 21 for HE staining to analyze differences in hair follicle cycling; transcriptome sequencing was performed on skin samples collected on day 4. Statistical analyses were conducted using the t test. Results:(1) Transmission electron microscopy showed that nanovesicles exhibited a spherical membranous structure with diameters of 141.3 ± 60.0 nm. (2) In 6 C57BL/6 female mice with full-thickness skin wounds, the wound area on day 12 was significantly smaller in the nanovesicle group (1.27 ± 0.50 mm 2) than in the control group (4.13 ± 1.03 mm 2, t = 4.34, P = 0.012). (3) Fluorescence microscopy revealed that nanovesicles were taken up by DPCs within 20 hours; the absorbance of DPCs was significantly higher in the nanovesicle group than in the control group ( t = 20.23, P < 0.001), and the percentage of EdU-positive cells was also significantly higher in the nanovesicle group (49.62% ± 6.45%) than in the control group (37.58% ± 3.42%, t = 3.69, P = 0.006). (4) In vivo imaging of the 6 C57BL/6 female mice showed strong fluorescence in the back of mice in the nanovesicle group on day 0, which markedly decreased by day 8, while no fluorescence was observed in the control group throughout the experiment. (5) Hair follicle cycle experiments on the 24 C57BL/6 female mice with depilated backs showed that the hair follicle length on day 4 after depilation was significantly longer in the nanovesicle group (368.00 ± 63.17 μm) than in the control group (266.90 ± 34.41 μm, t = 9.87, P < 0.001), and the hair bulb diameter was also significantly longer in the nanovesicle group (54.83 ± 10.32 μm) than in the control group (39.12 ± 7.54 μm, t = 16.02, P < 0.001) ; on day 18, the nanovesicle group showed a significantly higher hair follicle density (19.12 ± 0.90) compared with the control group (11.07 ± 1.51, t = 7.92, P = 0.001) ; on day 21, 46.13% ± 8.64% of hair follicles in the nanovesicle group remained in the anagen phase Ⅵ to the catagen phase Ⅱ, and 46.24% ± 3.29% were in the catagen phases Ⅲ to Ⅳ, while 78.89% ± 18.36% of hair follicles in the control group were in the telogen phases Ⅶ to Ⅷ. Transcriptome sequencing showed that differentially expressed genes in the nanovesicle group were significantly positively enriched in the keratinization process (NES = 2.23, P < 0.001) . Conclusion:Umbilical cord MSC-derived nanovesicles could promote the proliferation of DPCs, advance the entry of hair follicles into the anagen phase, delay their entry into the catagen phase, and induce hair regeneration.

Objective To examine the role and mechanism of clopidogrel in the development of salt-sensitive hypertension.Methods 8-week-old Dahl salt-sensitive(Dahl SS)rats and control salt-resistant(SS13BN)rats were divided randomly into six groups and fed for 8 weeks with normal salt(0.4％NaCl,NS),high salt(8％NaCl,HS),or high salt combined with clopidogrel gavage(8％NaCl+10 mg/(kg·d))clopidogrel,HS+CLO).Arterial systolic blood pressure was measured continuously over 8 weeks by the tail-cuff method,and systolic blood pressure was measured by carotid cannulation after 8 weeks(56 days).Renal histopathology was observed by hematoxylin and eosin staining,and renal inflammatory cell infiltration was detected by immunohistochemistry.Peripheral blood platelet activation and platelet-leukocyte aggregation were analyzed by flow cytometry,and the renal inflammation-related proteins tumor necrosis factor-α,interleukin(IL)-1β,IL-6,and key proteins in the p38MAPK/nuclear factor(NF)-κB signaling pathway were detected by Western blot.Results Compared with the NS group,Dahl SS HS rats had significantly increased blood pressure(P＜0.05),aggravated renal tissue damage,increased inflammatory cell infiltration,increased expression of inflammatory cytokines(P＜0.05),elevated peripheral blood platelet activation(P＜0.05)and platelet-leukocyte aggregation(P＜0.05),and increased expression of p38MAPK/NF-κB signaling pathway proteins(P＜0.05).Clopidogrel effectively alleviated these phenotypes induced by high salt in Dahl SS rats.Conclusions Clopidogrel alleviated high-salt-induced salt-sensitive hypertension and decreased renal inflammatory responses and dysfunction in Dahl SS rats by inhibiting platelet activation and the p38MAPK/NF-κB signaling pathway.

Objective:To investigate the effect of verbascoside(VERB)on high-fat diet-induced atherosclerosis(AS)in ApoE-/-mice and the effect on high mobility histone 1(HMGB1)/receptor for glycation end products(RAGE)/nuclear factor κB(NF-κB)pathway.Methods:A total of 90 ApoE-/-mice were randomly divided into normal group,AS group,VERB group,simvastatin group and VERB+pathway activator HMGB1 group,with 18 mice per group.After 8 weeks of group administration,blood and aorta samples were taken.Fasting serum triacylglycerol(TG),total cholesterol(TC)and low density lipoprotein(LDL)levels were deter-mined by automatic biochemical analyzer.Oil red O,HE and TUNEL staining were performed to observe apoptosis of aortic plaque and endothelial cell(EC).Flow cytometry was performed to analyze circulating EC numbers.Immunohistochemistry was performed to analyze the infiltration area of macrophages(CD68+)and T lymphocytes(CD3+)in aortic plaques.Western blot was performed to detect expressions of HMGB1/RAGE/NF-κB pathway,inflammation and adhesion molecules.Results:Compared with normal group,AS group had lipid plaques in arterial intima,thickness of the media was uneven,TG,TC,LDL levels,lession proportion,plaque area,circulating EC number,arterial EC apoptosis rate,macrophage(CD68+)and T lymphocyte(CD3+)infiltration areas,TNF-α,monocyte chemoattractant protein-1(MCP-1),vascular cell adhesion molecule-1(VCAM-1),intercellular adhesion molecule-1(ICAM-1),cytoplasm HMGB1,total HMGB1,total RAGE protein levels and nuclear/total p65 NF-κB levels were increased(P<0.05),while nuclear HMGB1 protein,nuclear/total HMGB1 and cytosolic p65 NF-κB levels were decreased(P<0.05).After VERB or simvastatin intervention,arterial lesions were alleviated,TG,TC,LDL levels,lession proportion,plaque area,circulating EC number,arterial EC apoptosis rate,macrophage(CD68+)and T lymphocyte(CD3+)infiltration areas,TNF-α,MCP-1,VCAM-1,ICAM-1,cytoplasm HMGB1,total HMGB1,RAGE protein levels and nuclear/total p65 NF-κB level were decreased(P<0.05),while nuclear HMGB1 protein,nuclear/total HMGB1 and cytosolic p65 NF-κB levels were increased(P<0.05),and HMGB1 was able to antagonize the protective effect of VERB on AS mice.Conclusion:VERB can inhibit expressions of inflammatory and adhesion fac-tors in arterial plaques in ApoE-/-mice,reduce EC shedding and apoptosis,therefore improve AS symptoms in ApoE-/-mice,and the mechanism may be related to the inhibition of HMGB1/RAGE and NF-κB pathway.

Objective:To explore the clinical and pathological features and survival outcomes of patients with early-onset intrahepatic cholangiocarcinoma (EOICC).Methods:This is a multicenter, retrospective cohort study. Data of 1 160 intrahepatic cholangiocarcinoma patients undergoing radical resection in 14 tertiary Grade A hospitals in China from January 2010 to November 2021 were retrospectively collected. The cohort included 632 males and 528 females, aged( M (IQR)) 61 (14) years (range: 22 to 93 years). ICC aged ≤50 years at the time of diagnosis was defined as EOICC and >50 years as late-onset intrahepatic cholangiocarcinoma (LOICC). Of these, there were 247 cases in the EOICC group and 913 cases in the LOICC. The clinical and pathological characteristics of both groups were analyzed and compared using the independent sample t-test, Mann-Whitney U test or Kaplan-Meier method. Univariate and multivariate Cox regression models for patient outcomes were constructed and forest graphed. Results:Compared with the patients in the LOICC group, patients in the EOICC group had lower carcinoembryonic antigen levels (2.5(4.0) μg/L vs. 3.1(5.2)μg/L, U=124 899, P=0.009) and CA19-9 level (63.4(524.7)U/ml vs. 77.9(611.3)U/ml, U=120 320, P=0.013), higher levels of ALT (29(35)U/L vs. 24(26)U/L, U=101 214, P=0.013), a lower score of the Eastern US Cooperative Oncology Group (0 score patients: 54.7% vs. 44.1%, χ2=12.472, P=0.014), higher TNM stage ( χ2=11.807, P=0.038), and proportion of lymph node dissection (62.3% vs. 54.1%, χ2=5.355, P=0.021). Patients in the two groups in sex, first diagnosis symptoms, intrahepatic bile duct stone history, nail protein, albumin, total bilirubin, transaminase, liver function Child-Pugh grade, T stage, stage, N stage, preoperative laparoscopic exploration proportion, tumor diameter, vascular invasion proportion, differentiation, margin, intraoperative bleeding, postoperative complications, postoperative hospital days were no statistical significance (all P>0.05). Patients in the EOICC group had better outcomes than the LOICC group (median survival time: 29.7 months vs. 25.0 months, 3-year overall survival: 45.1% vs. 37.8%, P=0.027). Conclusion:EOICC patients are better than LOICC patients in carcinoembryonic antigen, CA19-9, ALT, physical strength status and TNM stage, and the long-term prognosis is also better than LOICC patients.