Manufacturing, quality control, and GLP-grade preclinical study of nebulized allogenic adipose mesenchymal stromal cells-derived extracellular vesicles.

BACKGROUND: Human adipose stromal cells-derived extracellular vesicles (haMSC-EVs) have been shown to alleviate inflammation in acute lung injury (ALI) animal models. However, there are few systemic studies on clinical-grade haMSC-EVs. Our study aimed to investigate the manufacturing, quality control (QC) and preclinical safety of clinical-grade haMSC-EVs. METHODS: haMSC-EVs were isolated from the conditioned medium of human adipose MSCs incubated in 2D containers. Purification was performed by PEG precipitation and differential centrifugation. Characterizations were conducted by nanoparticle tracking analysis, transmission electron microscopy (TEM), Western blotting, nanoflow cytometry analysis, and the TNF-α inhibition ratio of macrophage [after stimulated by lipopolysaccharide (LPS)]. RNA-seq and proteomic analysis with liquid chromatography tandem mass spectrometry (LC-MS/MS) were used to inspect the lot-to-lot consistency of the EV products. Repeated toxicity was evaluated in rats after administration using trace liquid endotracheal nebulizers for 28 days, and respiratory toxicity was evaluated 24 h after the first administration. In vivo therapeutic effects were assessed in an LPS-induced ALI/ acute respiratory distress syndrome (ARDS) rat model. RESULTS: The quality criteria have been standardized. In a stability study, haMSC-EVs were found to remain stable after 6 months of storage at - 80°C, 3 months at - 20 °C, and 6 h at room temperature. The microRNA profile and proteome of haMSC-EVs demonstrated suitable lot-to-lot consistency, further suggesting the stability of the production processes. Intratracheally administered 1.5 × 108 particles/rat/day for four weeks elicited no significant toxicity in rats. In LPS-induced ALI/ARDS model rats, intratracheally administered haMSC-EVs alleviated lung injury, possibly by reducing the serum level of inflammatory factors. CONCLUSION: haMSC-EVs, as an off-shelf drug, have suitable stability and lot-to-lot consistency. Intratracheally administered haMSC-EVs demonstrated excellent safety at the tested dosages in systematic preclinical toxicity studies. Intratracheally administered haMSC-EVs improved the lung function and exerted anti-inflammatory effects on LPS-induced ALI/ARDS model rats.

Added value of arterial enhancement fraction derived from dual-energy computed tomography for preoperative diagnosis of cervical lymph node metastasis in papillary thyroid cancer: initial results.

OBJECTIVES: To explore the added value of arterial enhancement fraction (AEF) derived from dual-energy computed tomography CT (DECT) to conventional image features for diagnosing cervical lymph node (LN) metastasis in papillary thyroid cancer (PTC). METHODS: A total of 273 cervical LNs (153 non-metastatic and 120 metastatic) were recruited from 92 patients with PTC. Qualitative image features of LNs were assessed. Both single-energy CT (SECT)-derived AEF (AEFS) and DECT-derived AEF (AEFD) were calculated. Correlation between AEFD and AEFS was determined using Pearson's correlation coefficient. Multivariate logistic regression analysis with the forward variable selection method was used to build three models (conventional features, conventional features + AEFS, and conventional features + AEFD). Diagnostic performances were evaluated using receiver operating characteristic (ROC) curve analyses. RESULTS: Abnormal enhancement, calcification, and cystic change were chosen to build model 1 and the model provided moderate diagnostic performance with an area under the ROC curve (AUC) of 0.675. Metastatic LNs demonstrated both significantly higher AEFD (1.14 vs 0.48; p < 0.001) and AEFS (1.08 vs 0.38; p < 0.001) than non-metastatic LNs. AEFD correlated well with AEFS (r = 0.802; p < 0.001), and exhibited comparable performance with AEFS (AUC, 0.867 vs 0.852; p = 0.628). Combining CT image features with AEFS (model 2) and AEFD (model 3) could significantly improve diagnostic performances (AUC, 0.865 vs 0.675; AUC, 0.883 vs 0.675; both p < 0.001). CONCLUSIONS: AEFD correlated well with AEFS, and exhibited comparable performance with AEFS. Integrating qualitative CT image features with both AEFS and AEFD could further improve the ability in diagnosing cervical LN metastasis in PTC. CLINICAL RELEVANCE STATEMENT: Arterial enhancement fraction (AEF) values, especially AEF derived from dual-energy computed tomography, can help to diagnose cervical lymph node metastasis in patients with papillary thyroid cancer, and complement conventional CT image features for improved clinical decision making. KEY POINTS: • Metastatic cervical lymph nodes (LNs) demonstrated significantly higher arterial enhancement fraction (AEF) derived from dual-energy computed tomography (DECT) and single-energy CT (SECT)-derived AEF (AEFS) than non-metastatic LNs in patients with papillary thyroid cancer. • DECT-derived AEF (AEFD) correlated significantly with AEFS, and exhibited comparable performance with AEFS. • Integrating qualitative CT images features with both AEFS and AEFD could further improve the differential ability.

Right time to detect urine iodine during papillary thyroid carcinoma diagnosis and treatment: A case report.

BACKGROUND: This is the first documentation of a spontaneous and nonspecific chemical reaction of an iodinated contrast media with ammonium persulfate used in As3+-Ce4+ catalytic spectrophotometry for urine iodine concentration (UIC) detection. CASE SUMMARY: We herein report an incidental case who had a dual source computed tomography examination for papillary thyroid carcinoma diagnosis. Serial spot urine specimens were collected during her hospitalization and were measured by As3+-Ce4+ catalytic spectrophotometry on a Beckman Coulter AU5800. The reacted solutions were "brownish", and the results showed extremely high iodine concentrations despite serial dilutions. The patient claimed no dietary habit of iodized salt or iodine-containing medical history, which strongly pointed to iodinated contrast media (ICM) via intravenous injection. Even with 0.01% ICM, its interruption is still profound on the desired urine iodine reaction with ammonium persulfate, leading to inaccurate UIC and possibly inappropriate treatment. CONCLUSION: The following laboratory suggestions should be considered: (1) As3+-Ce4+ catalytic spectrophotometry is only suitable for UIC measurement after confirmed ICM renal clearance; (2) A mass spectrometry-based method can be applied as an alternative during the ICM clearance period; and (3) The UIC baseline can be confirmed after ICM injection by consecutive detection for at least 2 mo.

Extracellular Volume Fraction Derived From Dual-Layer Spectral Detector Computed Tomography for Diagnosing Cervical Lymph Nodes Metastasis in Patients With Papillary Thyroid Cancer: A Preliminary Study.

Objectives: The current study evaluates the performance of dual-energy computed tomography (DECT) derived extracellular volume (ECV) fraction based on dual-layer spectral detector CT for diagnosing cervical lymph nodes (LNs) metastasis from papillary thyroid cancer (PTC) and compares it with the value of ECV derived from conventional single-energy CT (SECT). Methods: One hundred and fifty-seven cervical LNs (81 non-metastatic and 76 metastatic) were recruited. Among them, 59 cervical LNs (27 non-metastatic and 32 metastatic) were affected by cervical root artifact on the contrast-enhanced CT images in the arterial phase. Both the SECT-derived ECV fraction (ECVS) and the DECT-derived ECV fraction (ECVD) were calculated. A Pearson correlation coefficient and a Bland-Altman analysis were performed to evaluate the correlations between ECVD and ECVS. Receiver operator characteristic curves analysis and the Delong method were performed to assess and compare the diagnostic performance. Results: ECVD correlated significantly with ECVS (r = 0.925; p <0.001) with a small bias (-0.6). Metastatic LNs showed significantly higher ECVD (42.41% vs 22.53%, p <0.001) and ECVS (39.18% vs 25.45%, p <0.001) than non-metastatic LNs. By setting an ECVD of 36.45% as the cut-off value, optimal diagnostic performance could be achieved (AUC = 0.813), which was comparable with that of ECVS (cut-off value = 34.99%; AUC = 0.793) (p = 0.265). For LNs affected by cervical root artifact, ECVD also showed favorable efficiency (AUC = 0.756), which was also comparable with that of ECVS (AUC = 0.716) (p = 0.244). Conclusions: ECVD showed a significant correlation with ECVS. Compared with ECVS, ECVD showed comparable performance in diagnosing metastatic cervical LNs in PTC patients, even though the LNs were affected by cervical root artifacts on arterial phase CT.

Nebulized exosomes derived from allogenic adipose tissue mesenchymal stromal cells in patients with severe COVID-19: a pilot study.

BACKGROUND: Existing clinical studies supported the potential efficacy of mesenchymal stromal cells as well as derived exosomes in the treatment of COVID-19. We aimed to explore the safety and efficiency of aerosol inhalation of the exosomes derived from human adipose-derived MSCs (haMSC-Exos) in patients with COVID-19. METHODS: The MEXCOVID trial is a phase 2a single-arm, open-labelled, interventional trial and patients were enrolled in Jinyintan Hospital, Wuhan, China. Eligible 7 patients were assigned to receive the daily dose of haMSCs-Exos (2.0 × 108 nano vesicles) for consecutively 5 days. The primary outcomes included the incidence of prespecified inhalation-associated events and serious adverse events. We also observed the demographic data, clinical characteristics, laboratory results including lymphocyte count, levels of D-dimer and IL-6 as well as chest imaging. RESULTS: Seven severe COVID-19 related pneumonia patients (4 males and 3 females) were enrolled and received nebulized haMSC-Exos. The median age was 57 year (interquartile range (IQR), 43 year to 70 year). The median time from onset of symptoms to hospital admission and administration of nebulized haMSC-Exos was 30 days (IQR, 15 days to 40 days) and 54 d (IQR, 34 d to 69 d), respectively. All COVID-19 patients tolerated the haMSC-Exos nebulization well, with no evidence of prespecified adverse events or clinical instability during the nebulization or during the immediate post-nebulization period. All patients presented a slight increase of serum lymphocyte counts (median as 1.61 × 109/L vs. 1.78 × 109/L). Different degrees of resolution of pulmonary lesions after aerosol inhalation of haMSC-Exos were observed among all patients, more obviously in 4 of 7 patients. CONCLUSIONS: Our trial shows that a consecutive 5 days inhalation dose of clinical grade haMSC-Exos up to a total amount of 2.0 × 109 nano vesicles was feasible and well tolerated in seven COVID-19 patients, with no evidence of prespecified adverse events, immediate clinical instability, or dose-relevant toxicity at any of the doses tested. This safety profile is seemingly followed by CT imaging improvement within 7 days. Further trials will have to confirm the long-term safety or efficacy in larger population. TRIAL REGISTRATION: MEXCOVID, NCT04276987.

Radiomics from Primary Tumor on Dual-Energy CT Derived Iodine Maps can Predict Cervical Lymph Node Metastasis in Papillary Thyroid Cancer.

RATIONALE AND OBJECTIVES: To develop and validate 2 iodine maps based radiomics nomograms for preoperatively predicting cervical lymph node metastasis (LNM) and central lymph node metastasis (CLNM) in papillary thyroid cancer (PTC). MATERIALS AND METHODS: A total of 346 patients with PTC were enrolled and allocated to training (242) and validation (104) sets. Radiomics features were extracted from arterial and venous phase iodine maps, respectively. Aggregated machine-learning strategy was applied for features selection and construction of 2 radiomics scores (LN rad-score; CLN rad-score). Logistic regression model was employed to establish two radiomics nomograms (nomogram 1: predicting LNM; nomogram 2: predicting CLNM) after incorporating LN or CLN rad-score with clinical predictors. Nomograms performance was determined by discrimination, calibration and clinical usefulness. RESULTS: Nomogram 1 incorporated LN rad-score, age (categorized by 55) and CT reported LN status; Nomogram 2 incorporated CLN rad-score, capsule contact >25% and CT reported CLN status. 2 nomograms both showed good discrimination and calibration in the training (AUC = 0.847; AUC = 0.837) and validation cohorts (AUC = 0.807; AUC = 0.795). Significant improved AUC, net reclassification index (NRI) and integrated discriminatory improvement (IDI) confirmed additional great predictive value of 2 rad-scores, compared with clinical models without radiomics. Decision curve analysis indicated clinical utility of nomograms. 2 nomograms both demonstrated favorable predictive efficacy in CT reported LN or CLN negative subgroup (AUC = 0.766; AUC = 0.744). CONCLUSION: The presented 2 radiomics nomograms are useful tools for preoperative prediction of LNM and CLNM in PTC.

Feasibility study of using virtual non-contrast images derived from dual-energy CT to replace true non-contrast images in patients diagnosed with papillary thyroid carcinoma.

OBJECTIVE: To assess the feasibility of using virtual non-contrast (VNC) images derived from dual-energy computed tomography (DECT) to replace true non-contrast (TNC) images of papillary thyroid carcinoma (PTC) patients. METHODS: Images of 96 PTC patients were retrospectively analyzed. TNC images were acquired under the single-energy mode of DECT after the plain scanning. The arterial and venous phase VNC (VNC-a and VNC-v) images were generated by the post-processing algorithm from the arterial phase and venous phase of contrast-enhanced CT images, respectively. Mean attenuation values, image noise, number and length of calcification were measured. Radiation dose was also calculated. Last, subjective score of image quality was evaluated by a 5-point scale. RESULTS: Signal-to-noise ratio (SNR) of each tissue in TNC images is significantly higher than that of VNC images (p<0.050). Contrast-to-noise ratio (CNR) of fat, muscle, thyroid nodules and internal carotid artery in TNC images is significantly higher than that of VNC images, while CNR in TNC images is lower for cervical vertebra (p<0.001). Calcification is detected on TNC images of 44 patients, while it is omitted on VNC images of 14 patients (31.8%). The subjective score of TNC images is higher than VNC images (p<0.001). The effective dose reduction is 47.6% by avoiding plain scanning. CONCLUSIONS: Considering the different attenuation value, SNR, CNR and especially reduced detection rate of calcification, we deem that VNC images cannot be directly used to replace TNC images in PTC patients, despite the reduced radiation dose.

Management of Sonographically Suspicious Thyroid Nodules 1 cm or Smaller and Candidacy for Active Surveillance: Experience of a Tertiary Center in China.

OBJECTIVE: Our objective was to investigate the management of patients with asymptomatic suspicious thyroid nodules ≤1 cm. METHODS: We retrospectively reviewed medical records of patients with sonographically suspicious thyroid nodules ≤1 cm and without distant metastases, suspicious lymph node metastasis (LNM), or extrathyroidal extension (ETE). RESULTS: Of the 386 enrolled patients, 174 (45.1%) had immediate surgery (IS), while 212 (54.9%) underwent active surveillance (AS). In the IS group, 166 (95.4%) patients were confirmed as having papillary thyroid microcarcinoma. LNM and ETE were observed in 24.7% and 2.4% cases, respectively. In the AS group, nodule size increased by ≥3 mm in 11 (5.2%) patients and 39 (18.4%) had a >50% increase in nodule volume after a median follow-up of 12 months. Nodules with smaller volume at diagnosis were more likely to increase in volume later. Newly suspicious LNM was detected in 23 (10.8%) patients. Delayed surgery (DS) was performed in 101 patients, with 27 showing disease progression. ETE and LNM were detected in 3% and 36%, respectively, of patients with papillary thyroid microcarcinoma. Compared with IS, tumors in the DS group more frequently showed lateral LNM and capsular invasion (P < .05). No patient had recurrence or died of thyroid cancer during postoperative follow-up (median 26 [4-60] months). CONCLUSIONS: IS or DS of patients with asymptomatic suspicious thyroid nodules ≤1 cm was relatively high in China. The inertia of low-risk nodules and the effectiveness of DS for those that progressed make AS a feasible strategy.

Texture analysis of dual-phase contrast-enhanced CT in the diagnosis of cervical lymph node metastasis in patients with papillary thyroid cancer.

BACKGROUND: Computed tomography texture analysis (CTTA) provides objective and quantitative information regarding tumor heterogeneity beyond visual inspection. However, no study has yet used CTTA to differentiate metastatic from non-metastatic cervical lymph node in patients with papillary thyroid cancer (PTC). PURPOSE: To evaluate the value of texture analysis of dual-phase contrast-enhanced CT images in diagnosing cervical lymph node metastasis in patients with PTC. MATERIAL AND METHODS: Metastatic (n = 27) and non-metastatic (n = 32) cervical lymph nodes were analyzed retrospectively. Texture analyses were performed on both arterial (A) and venous (V) phase CT images. Texture parameters, including mean gray-level intensity, skewness, kurtosis, entropy, and uniformity, were obtained and compared between groups. Receiver operating characteristic (ROC) curves analyses and multivariate logistic regression analysis were used in our study. RESULTS: Metastatic lymph nodes showed significantly higher A-mean gray-level intensity, A-entropy, and lower A-kurtosis and V-kurtosis (all P < 0.001) than non-metastatic mimics. The ROC curve analyses indicated that A-kurtosis demonstrated an optimal diagnostic area under the curve (AUC; 0.884) and specificity (92.59%), while the A-mean gray-level intensity showed optimal diagnostic sensitivity (90.62%). Multivariate logistic regression analysis showed that A-mean gray-level intensity (P = 0.006, odds ratio [OR] = 24.297) and V-kurtosis (P = 0.014, OR = 19.651) were the independent predictor for metastatic cervical lymph node. CONCLUSION: Dual-phase contrast-enhanced CCTA-especially A-mean gray-level intensity and V-kurtosis-may have the potential to diagnose metastatic cervical lymph node in patients with PTC.

Radiomics analysis of dual-energy CT-derived iodine maps for diagnosing metastatic cervical lymph nodes in patients with papillary thyroid cancer.

OBJECTIVE: To investigate the value of radiomics analysis of dual-energy computed tomography (DECT)-derived iodine maps for preoperative diagnosing cervical lymph nodes (LNs) metastasis in patients with papillary thyroid cancer (PTC). METHODS: Two hundred and fifty-five LNs (143 non-metastatic and 112 metastatic) were enrolled and allocated to training and validation sets (7:3 ratio). Radiomics features were extracted from arterial and venous phase iodine maps, respectively. Radiomics signature was constructed based on reproducible features using the least absolute shrinkage and selection operator (LASSO) logistic regression algorithm with 10-fold cross-validation. Logistic regression modeling was employed to build models based on CT image features (model 1), radiomics signature (model 2), and the combined (model 3). A nomogram was plotted for the combined model and decision curve analysis was applied for clinical use. Diagnostic performance was assessed and compared. Internal validation was performed on an independent set containing 78 LNs. RESULTS: Model 3 showed optimal diagnostic performance in both training (AUC = 0.933) and validation set (AUC = 0.895), followed by model 2 (training set, AUC = 0.910; validation set, AUC = 0.847). Both these two models outperformed model 1 in both training (AUC = 0.763) (p < 0.05) and validation set (AUC = 0.728) (p < 0.05). CONCLUSION: Radiomics analysis of DECT-derived iodine maps showed better diagnostic performance than qualitative evaluation of CT image features in preoperative diagnosing cervical LN metastasis in PTC patients. Radiomics signature integrated with CT image features can serve as a promising imaging biomarker for the differentiation. KEY POINTS: • Conventional CT image features have limited value for the diagnosis of metastatic LNs in PTC patients. • Radiomics analysis of dual-energy CT-derived iodine maps significantly outperformed qualitative CT image features in differentiating metastatic from non-metastatic LNs. • Radiomics signature integrated with qualitative CT image features can serve as a useful tool in judging LNs status, thus aiding clinical decision-making.

Application of high-resolution ultrasound, real-time elastography, and contrast-enhanced ultrasound in differentiating solid thyroid nodules.

High-resolution ultrasound (HRUS) is a sensitive tool for identifying thyroid nodules. Real-time elastography (RTE) and contrast-enhanced ultrasound (CEUS) are newly developed methods which could measure tissue elasticity and perfusion features. The aim of the present study was to evaluate and compare the diagnostic efficiency of HRUS, RTE, CEUS and their combined use in the differentiation of benign and malignant solid thyroid nodules.In total, 111 consecutive patients with 145 thyroid nodules who were scheduled for surgery were included in the study. All of them underwent HRUS, RTE, and CEUS examination. The independent ultrasound (US) predictors for malignancy were determined and quantified using logistic regression analysis, based on which a risk-scoring model was established for each method. The diagnostic efficiency of each method was assessed by receiver operating characteristic (ROC) curve analysis.HRUS showed the best diagnostic efficiency among the 3 US methods, with 74.6% sensitivity and 87.8% specificity. CEUS had higher sensitivity (85.7%), whereas RTE alone did not show much advantage. Combined use of RTE and HRUS increased the sensitivity (92.1%). The HRUS-RTE-CEUS combination could increase both the sensitivity and specificity (87.3%, 91.5%), with the best AUC (0.935) among all the methods.The overall diagnostic value of HRUS in predicting malignancy is the best among the 3 US methods. Combined use of RTE and CEUS and HRUS could improve the diagnostic efficiency for solid thyroid nodules.

Value of TIRADS, BSRTC and FNA-BRAF V600E mutation analysis in differentiating high-risk thyroid nodules.

The thyroid imaging reporting and data system (TIRADS) and Bethesda system for reporting thyroid cytopathology (BSRTC) have been used for interpretation of ultrasound and fine-needle aspiration cytology (FNAC) results of thyroid nodules. BRAF(V600E) mutation analysis is a molecular tool in diagnosing thyroid carcinoma. Our objective was to compare the diagnostic value of these methods in differentiating high-risk thyroid nodules. Total 220 patients with high-risk thyroid nodules were recruited in this prospective study. They all underwent ultrasound, FNAC and BRAF(V600E) mutation analysis. The sensitivity and specificity of TIRADS were 73.1% and 88.4%. BSRTC had higher specificity (97.7%) and similar sensitivity (77.6%) compared with TIRADS. The sensitivity and specificity of BRAF(V600E) mutation (85.1%, 100%) were the highest. The combination of BSRTC and BRAF(V600E) mutation analysis significantly increased the efficiency, with 97.8% sensitivity, 97.7% specificity. In patients with BSRTC I-III, the mutation rate of BRAF(V600E) was 64.5% in nodules with TIRADS 4B compared with 8.4% in nodules with TIRADS 3 or 4A (P < 0.001). Our study indicated that combination of BSRTC and BRAF(V600E) mutation analysis bears a great value in differentiating high-risk thyroid nodules. The TIRADS is useful in selecting high-risk patients for FNAB and patients with BSRTC I-III for BRAF(V600E) mutation analysis.

Annual financial impact of thyroidectomies for nodular thyroid disease in China.

A large proportion of patients with thyroid nodules in China undergo thyroidectomy in order to get confirmatory histology diagnosis. The financial impact of this modality remains to be investigated. To evaluate rationality of performing thyroidectomy without a routine FNA preoperatively from the economic perspective, we conducted a retrospective, observational study of all archival thyroidectomies with records of cost per stay (CPS), cost per day (CPD) and length of stay (LOS) from 2008 to 2013 in the First Affiliated Hospital of Nanjing Medical University. We compared all the parameters between cancer and non-cancer thyroidectomies. We recruited 6, 140 thyroidectomies with valid records of CPS, CPD and LOS in this period. The CPS of cancer thyroidectomy was significantly higher than non-cancer thyroidectomy. The percentage of cancer thyroidectomy increased from 26.5% to 41.6%. The percentage of annual cost of cancer thyroidectomies rose from 30.2% to 45.2%. The LOS for cancer and non-cancer thyroidectomy decreased while the CPD increased in the past six years. The estimated national cost in 2012 for all thyroidectomies would be USD 1.86 billion with USD 1.09 billion for non-cancer thyroidectomies. We have witnessed great improvement in the healthcare for patients with thyroid nodules in China. However, given limited healthcare resources, currently thyroid FNA for more precise preoperative diagnosis may help to curb the rapidly increasing demand in healthcare costs in the future for nodular thyroid disease in China.