CT texture analysis in predicting treatment response and survival in patients with hepatocellular carcinoma treated with transarterial chemoembolization using random forest models.

BACKGROUND: Using texture features derived from contrast-enhanced computed tomography (CT) combined with general imaging features as well as clinical information to predict treatment response and survival in patients with hepatocellular carcinoma (HCC) who received transarterial chemoembolization (TACE) treatment. METHODS: From January 2014 to November 2022, 289 patients with HCC who underwent TACE were retrospectively reviewed. Their clinical information was documented. Their treatment-naïve contrast-enhanced CTs were retrieved and reviewed by two independent radiologists. Four general imaging features were evaluated. Texture features were extracted based on the regions of interest (ROIs) drawn on the slice with the largest axial diameter of all lesions using Pyradiomics v3.0.1. After excluding features with low reproducibility and low predictive value, the remaining features were selected for further analyses. The data were randomly divided in a ratio of 8:2 for model training and testing. Random forest classifiers were built to predict patient response to TACE treatment. Random survival forest models were constructed to predict overall survival (OS) and progress-free survival (PFS). RESULTS: We retrospectively evaluated 289 patients (55.4 ± 12.4 years old) with HCC treated with TACE. Twenty features, including 2 clinical features (ALT and AFP levels), 1 general imaging feature (presence or absence of portal vein thrombus) and 17 texture features, were included in model construction. The random forest classifier achieved an area under the curve (AUC) of 0.947 with an accuracy of 89.5% for predicting treatment response. The random survival forest showed good predictive performance with out-of-bag error rate of 0.347 (0.374) and a continuous ranked probability score (CRPS) of 0.170 (0.067) for the prediction of OS (PFS). CONCLUSIONS: Random forest algorithm based on texture features combined with general imaging features and clinical information is a robust method for predicting prognosis in patients with HCC treated with TACE, which may help avoid additional examinations and assist in treatment planning.

Association between High Diffusion-Weighted Imaging-Derived Functional Tumor Burden of Peritoneal Carcinomatosis and Overall Survival in Patients with Advanced Ovarian Carcinoma.

OBJECTIVE: To investigate the association between functional tumor burden of peritoneal carcinomatosis (PC) derived from diffusion-weighted imaging (DWI) and overall survival in patients with advanced ovarian carcinoma (OC). MATERIALS AND METHODS: This prospective study was approved by the local research ethics committee, and informed consent was obtained. Fifty patients (mean age ± standard deviation, 57 ± 12 years) with stage III-IV OC scheduled for primary or interval debulking surgery (IDS) were recruited between June 2016 and December 2021. DWI (b values: 0, 400, and 800 s/mm²) was acquired with a 16-channel phased-array torso coil. The functional PC burden on DWI was derived based on K-means clustering to discard fat, air, and normal tissue. A score similar to the surgical peritoneal cancer index was assigned to each abdominopelvic region, with additional scores assigned to the involvement of critical sites, denoted as the functional peritoneal cancer index (fPCI). The apparent diffusion coefficient (ADC) of the largest lesion was calculated. Patients were dichotomized by immediate surgical outcome into high- and low-risk groups (with and without residual disease, respectively) with subsequent survival analysis using the Kaplan-Meier curve and log-rank test. Multivariable Cox proportional hazards regression was used to evaluate the association between DWI-derived results and overall survival. RESULTS: Fifteen (30.0%) patients underwent primary debulking surgery, and 35 (70.0%) patients received neoadjuvant chemotherapy followed by IDS. Complete tumor debulking was achieved in 32 patients. Patients with residual disease after debulking surgery had reduced overall survival (p = 0.043). The fPCI/ADC was negatively associated with overall survival when accounted for clinicopathological information with a hazard ratio of 1.254 for high fPCI/ADC (95% confidence interval, 1.007-1.560; p = 0.043). CONCLUSION: A high DWI-derived functional tumor burden was associated with decreased overall survival in patients with advanced OC.

CT texture analysis in histological classification of epithelial ovarian carcinoma.

OBJECTIVES: The study aimed to compare the ability of morphological and texture features derived from contrast-enhanced CT in histological subtyping of epithelial ovarian carcinoma (EOC). METHODS: Consecutive 205 patients with newly diagnosed EOC who underwent contrast-enhanced CT were included and dichotomised into high-grade serous carcinoma (HGSC) and non-HGSC. Clinical information including age and cancer antigen 125 (CA-125) was documented. The pre-treatment images were analysed using commercial software, TexRAD, by two independent radiologists. Eight qualitative CT morphological features were evaluated, and 36 CT texture features at 6 spatial scale factors (SSFs) were extracted per patient. Features' reduction was based on kappa score, intra-class correlation coefficient (ICC), univariate ROC analysis and Pearson's correlation test. Texture features with ICC ≥ 0.8 were compared by histological subtypes. Patients were randomly divided into training and testing sets by 8:2. Two random forest classifiers were determined and compared: model 1 incorporating selected morphological and clinical features and model 2 incorporating selected texture and clinical features. RESULTS: HGSC showed specifically higher texture features than non-HGSC (p < 0.05). Both models performed highly in predicting histological subtypes of EOC (model 1: AUC 0.891 and model 2: AUC 0.937), and no statistical significance was found between the two models (p = 0.464). CONCLUSION: CT texture analysis provides objective and quantitative metrics on tumour characteristics with HGSC demonstrating specifically high texture features. The model incorporating texture analysis could classify histology subtypes of EOC with high accuracy and performed as well as morphological features. KEY POINTS: • A number of CT morphological and texture features showed good inter- and intra-observer agreements. • High-grade serous ovarian carcinoma showed specifically higher CT texture features than non-high-grade serous ovarian carcinoma. • CT texture analysis could differentiate histological subtypes of epithelial ovarian carcinoma with high accuracy.

Molecular Imaging of Peritoneal Carcinomatosis in Ovarian Carcinoma.

OBJECTIVE. This article discusses the emerging roles of 18F-FDG PET/CT and DWI in the assessment of peritoneal carcinomatosis in ovarian carcinoma from diagnostic accuracy to disease prognostication with gross pathologic correlation. CONCLUSION. PET/CT and DWI have incremental clinical values over conventional modalities with high predictive values of incomplete cytoreduction in ovarian carcinoma. The respective quantitative metrics offer evaluation of tumor burden with prognostic value in ovarian carcinoma.

Functional tumour burden of peritoneal carcinomatosis derived from DWI could predict incomplete tumour debulking in advanced ovarian carcinoma.

OBJECTIVES: To investigate the predictive value of peritoneal carcinomatosis (PC) quantification by DWI in determining incomplete tumour debulking in ovarian carcinoma (OC). METHODS: Prospective patients with suspected stage III-IV or recurrent OC were recruited for DWI before surgery. PC on DWI was segmented semi-automatically by k-means clustering, retaining voxels with intermediate apparent diffusion coefficient (ADC) to quantify PC burden. A scoring system, functional peritoneal cancer index (fPCI), was proposed based on the segmentation of tumour volume in 13 abdominopelvic regions with additional point given to involvement of critical sites. ADC of the largest PC was recorded. The surgical complexity and outcomes (complete vs. incomplete tumour debulking) were documented. fPCI was correlated with surgical PCI (sPCI), surgical complexity, and its ability to predict incomplete tumour debulking. RESULTS: Fifty-three patients with stage III-IV or recurrent OC were included with a mean age of 56.1 ± 11.8 years old. Complete tumour debulking was achieved in 38/53 patients (71.7%). Significant correlation was found between fPCI and sPCI (r > 0.757, p < 0.001). Patients with high-fPCI (fPCI ≥ 6) had a high surgical complexity score (p = 0.043) with 84.2% received radical or supra-radical surgery. The mean fPCI was significantly higher in patients with incomplete tumour debulking than in those with complete debulking (10.27 vs. 4.71, p < 0.001). fPCI/ADC combined with The International Federation of Gynecology and Obstetrics stage achieved 92.5% accuracy in predicting incomplete tumour debulking (AUC 0.947). CONCLUSIONS: DWI-derived fPCI offered a semi-automated estimation of PC burden. fPCI/ADC could predict the likelihood of incomplete tumour debulking with high accuracy. KEY POINTS: • Functional peritoneal cancer index (fPCI) derived from DWI offered a semi-automated estimation of tumour burden in ovarian carcinoma. • fPCI was highly correlated with surgical PCI (sPCI). • fPCI/ADC could predict the likelihood of incomplete tumour debulking with high accuracy.

The Value of Contrast-Enhanced CT in the Detection of Residual Disease After Neo-Adjuvant Chemotherapy in Ovarian Cancer.

RATIONALE AND OBJECTIVES: To evaluate the diagnostic performance of contrast-enhanced computed tomography (CT) in predicting residual disease following neo-adjuvant chemotherapy (NACT) in stage III/IV ovarian cancer. MATERIALS AND METHODS: This was a retrospective observational cohort study including consecutive patients with primary stage III/IV ovarian cancer who received NACT before interval debulking surgery. CT findings before interval debulking surgerywere correlated with histological/surgical findings. Diagnostic characteristics were calculated on patient-based and lesion-based analyses. False negative results on peritoneal carcinomatosis detection were correlated with lesion size and site. RESULTS: On patient-based analysis, CT (n = 58) had a sensitivity, specificity, positive predictive value, negative predictive value and accuracy of 92.16%, 57.14%, 94.00%, 50.00%, and 87.93%. On lesion-based analysis, the sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were 63.01%, 73.47%, 82.51%, 50.00%, and 66.51%. False negative results were associated with lesion size (p < 0.001). The diagnostic performance of CT on the detection of peritoneal carcinomatosis was low at the subdiaphragmatic spaces, bowel serosa and mesentery (p < 0.001). CONCLUSION: CT had low negative predictive value in determining residual disease following NACT on both patient-based and lesion-based analyses, especially for non-measurable lesions and at the subdiaphragmatic spaces, bowel serosa and mesentery.

Qualitative and quantitative comparison of image quality between single-shot echo-planar and interleaved multi-shot echo-planar diffusion-weighted imaging in female pelvis.

OBJECTIVES: To qualitatively and quantitatively compare the image quality between single-shot echo-planar (SS-EPI) and multi-shot echo-planar (IMS-EPI) diffusion-weighted imaging (DWI) in female pelvis METHODS: This was a prospective study involving 80 females who underwent 3.0T pelvic magnetic resonance imaging (MRI). SS-EPI and IMS-EPI DWI were acquired with 3 b values (0, 400, 800 s/mm2). Two independent reviewers assessed the overall image quality, artifacts, sharpness, and lesion conspicuity based on a 5-point Likert scale. Regions of interest (ROI) were placed on the endometrium and the gluteus muscles to quantify the signal intensities and apparent diffusion coefficient (ADC). Signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), and geometric distortion were quantified on both sequences. Inter-rater agreement was assessed using κ statistics and Kendall test. Qualitative scores were compared using Wilcoxon signed-rank test and quantitative parameters were compared with paired t test and Bland-Altman analysis. RESULTS: IMS-EPI demonstrated better image quality than SS-EPI for all aspects evaluated (SS-EPI vs. IMS-EPI: overall quality 3.04 vs. 4.17, artifacts 3.09 vs. 3.99, sharpness 2.40 vs. 4.32, lesion conspicuity 3.20 vs. 4.25; p < 0.001). Good agreement and correlation were observed between two reviewers (SS-EPI κ 0.699, r 0.742; IMS-EPI κ 0.702, r 0.789). IMS-EPI showed lower geometric distortion, SNR, and CNR than SS-EPI (p < 0.050). There was no significant difference in the mean ADC between the two sequences. CONCLUSION: IMS-EPI showed better image quality with lower geometric distortion without affecting the quantification of ADC, though the SNR and CNR decreased due to post-processing limitations. KEY POINTS: • IMS-EPI showed better image quality than SS-EPI. • IMS-EPI showed lower geometric distortion without affecting ADC compared with SS-EPI. • The SNR and CNR of IMS-EPI decreased due to post-processing limitations.

Impact of region of interest size on transcranial sonography based computer-aided diagnosis for Parkinson's disease.

Transcranial sonography (TCS) has gained increasing application for diagnosis of Parkinson's disease (PD) in clinical practice in recent years, because most PD patients, even in the early stage of PD, have abnormal hyperechogenicity of the substantia nigra (SN) in brainstem shown in TCS images. Therefore, the region of interest (ROI) for feature extraction should cover the SN region in a computer-aided diagnosis (CAD) system. The ROI size naturally affects the feature representation. However, there currently exist no unified standard for determining the size of ROI. In this work, we quantitatively compare the performance of TCS-based CAD with three sizes of ROIs, namely the entire midbrain (EM) region, the half of midbrain (HoM) region and the SN region. The experimental results on the original extracted features and the features by dimensionality reduction show that ROI covering the EM region achieves the overall best diagnosis performance. The results indicates that the neighboring regions around SN might also have abnormal symptoms, which cannot be clearly observed with naked eyes. It suggests that the large ROI includes more information for feature representation to improve the diagnosis performance of TCS-based CAD for PD.

Progress on the Photocatalytic Reduction Removal of Chromium Contamination.

Rapid industrialization leads to increased wastewater discharge encompassing hexavalent chromium (Cr(VI)), which leads to serious environmental problems of toxicity and potential carcinogenicity. Removal of these species is normally carried out by ion-exchange, precipitation, membrane filtration, sorption, photocatalytic reduction, etc. This review mainly focuses on the photocatalytic and photoelectrocatalytic (PEC) reduction of Cr (VI), because of their advantages over other methods such as reduced risk of secondary pollution by non-reduced Cr (VI), no sludge formation, no need for a large amount of chemical reagents, clean and easy installation. The main factors influencing the photocatalytic reduction efficiency of Cr (VI) such as catalyst activity, solution pH, Cr adsorption on the catalyst and additives, are briefly discussed. Finally, a special emphasis is provided to the photoelectrocatalytic (PEC) reduction of Cr (VI).

Proteomic analysis and comparison of intra­ and extracranial cerebral atherosclerosis responses to hyperlipidemia in rabbits.

The present study aimed to investigate protein expression levels of intra­ and extracranial atherosclerosis in rabbits following administration of a high­fat diet. Rabbits were randomly divided into control (group A; n=9) and high­fat diet (group B; n=9) groups. At week 12, tissues were sectioned from the common carotid artery (CCA) and middle cerebral artery (MCA). Pathological analysis was performed. Differential protein expression levels were examined by 2­D gel electrophoresis (2­DE) and mass spectrometry (MS) analysis and validated by western blotting. Serum lipid levels, the intima­media thickness (IMT) and degree of atherosclerosis of the CCA and MCA were increased at week 12 in the high­fat diet group compared with rabbits that received a normal diet. 2­DE and MS analysis of the protein extracted from CCA and MCA detected >439 different proteins; the expression of 25 proteins was altered, and 8 proteins [albumin A chain, tropomyosin α­1 chain (TPM1), heat shock protein 70 (HSP70), α­smooth muscle actin, ß­galactose binding agglutinin, TPM4 isoform 2, cell keratin 9, single octylic acid glyceride ß­2) demonstrated significant alterations in expression levels. Due to limited antibody sources, only three differentially expressed proteins (TPM1, HSP70 and α­smooth muscle actin) were examined by western blotting. The results of our previous study demonstrated that hyperlipidemia affected the IMT of intracranial and extracranial cerebral arteries. In the present study, protein expression levels of TPM1 and α­smooth muscle actin from extracranial cerebral arteries were significantly increased compared with intracranial cerebral arteries; however, protein expression levels of HSP70 from intracranial cerebral arteries was increased compared with extracranial cerebral arteries. The differences may be closely associated with cell proliferation and metastasis, and oxidoreduction, in intra­ and extracranial cerebral atherosclerosis. HSP70 may have protective properties against atherosclerosis via underlying anti­inflammatory mechanisms, furthermore, differential protein expression levels (TPM1, HSP70 and α­smooth muscle actin) between intra­ and extracranial cerebral arteries may facilitate the identification of novel biological markers for the diagnosis and treatment of cerebral arteriosclerosis.

Defect-induced ripening of zeolitic-imidazolate framework ZIF-8 and its implication to vapor-phase membrane synthesis.

We report for the first time that ZIF-8 crystals undergo an Ostwald-ripening-like process without degradation in the presence of a ligand vapor. The ripening process is dependent on the defect density of the crystals: the more defective the more amenable to the ripening. The process was adapted to synthesize ultra-thin ZIF-8 membranes by vapor-phase secondary growth.

Magnetic Resonance Elastography for the Evaluation of Liver Fibrosis in Chronic Hepatitis B and C by Using Both Gradient-Recalled Echo and Spin-Echo Echo Planar Imaging: A Prospective Study.

OBJECTIVES: Magnetic resonance elastography (MRE) with three-dimensional spin-echo echo planar imaging (3D-SE-EPI) is a newly emerging noninvasive method for assessing liver fibrosis. We hypothesized that 3D-SE-EPI might have better diagnostic accuracy than conventional two-dimensional gradient-recalled echo (2D-GRE). METHODS: We prospectively included 179 consecutive patients with chronic hepatitis B (CHB) or C (CHC) who underwent both MRE and liver biopsy. Liver stiffness was measured by both 3D-SE-EPI and 2D-GRE for staging biopsy-proven liver fibrosis (using METAVIR scores). A receiver-operating characteristic analysis using the area under the receiver-operating characteristic curve (AUC) was used to compare the diagnostic performance in predicting liver fibrosis between these two techniques, and compared them to serum markers of fibrosis. RESULTS: The technical failure rate of 3D-SE-EPI (2.2%, n=4/179) was lower compared with 2D-GRE (8.3%, n=15/179). The stiffness measured by 3D-SE-EPI was slightly lower compared with 2D-GRE, with the mean difference of 0.57 kPa (Bland and Altman plot, 95% limits of agreement: -0.32 and 1.45 kPa). AUCs for the characterization of ≥F1, ≥F2, ≥F3, and F4 were 0.957 (95% confidence interval (CI): 0.913-0.983), 0.971 (0.932-0.991), 0.991 (0.961-0.999), and 0.979 (0.942-0.995) for 3D-SE-EPI, which was slightly higher compared with the AUCs for 2D-GRE at each fibrosis stage (0.948 (0.901-0.977), 0.959 (0.915-0.981), 0.979 (0.943-0.995), and 0.976 (0.938-0.994), respectively), although none reached statistical significance (P=0.160-0.585). In an "intention-to-diagnose" analysis, the diagnostic accuracy (the proportion of well-classified patients) by EPI (86.7-91.3%, n=169) was higher compared with GRE (80.9-82.1%, n=158) after applying optimal cutoffs. Both 3D-SE-EPI and 2D-GRE performed better than serum fibrosis markers. CONCLUSIONS: With respect to 2D-GRE, 3D-SE-EPI has the advantage of lower failure rate with equivalent high diagnostic performance for staging liver fibrosis in CHB/CHC patients, and thus more helpful for those challenging cases in 2D-GRE.

Heteroepitaxially grown zeolitic imidazolate framework membranes with unprecedented propylene/propane separation performances.

Propylene/propane separation is one of the most challenging separations, currently achieved by energy-intensive cryogenic distillation. Despite the great potential for energy-efficient membrane-based separations, no commercial membranes are currently available due to the limitations of current polymeric materials. Zeolitic imidazolate framework, ZIF-8, with the effective aperture size of ∼4.0 Å, has been shown to be very promising for propylene/propane separation. Despite the extensive research on ZIF-8 membranes, only a few reported ZIF-8 membranes have displayed good propylene/propane separation performances presumably due to the challenges of controlling the microstructures of polycrystalline membranes. Here we report the first well-intergrown membranes of ZIF-67 (Co-substituted ZIF-8) by heteroepitaxially growing ZIF-67 on ZIF-8 seed layers. The ZIF-67 membranes exhibited impressively high propylene/propane separation capabilities. Furthermore, when a tertiary growth of ZIF-8 layers was applied to heteroepitaxially grown ZIF-67 membranes, the membranes exhibited unprecedentedly high propylene/propane separation factors of ∼200 possibly due to enhanced grain boundary structure.

[Jinghuaweikang capsules combined with triple therapy in the treatment of Helicobacter pylori associated gastritis and duodenal ulcer and analysis of antibiotic resistance: a multicenter, randomized, controlled, clinical study].

OBJECTIVE: To explore the efficacy of Jinghuaweikang capsules plus triple therapy (LACJ) in treatment of Helicobacter pylori (H. pylori) associated gastritis or duodenal ulcer, compare it with bismuth-containing quadruple therapy (LACB) and standard triple therapy (LAC) and analyze the antibiotic sensitivity of gastric mucosal H. pylori strains from the failed patients. METHODS: A total of 565 patients with H. pylori infection were recruited from 11 hospitals from January 2010 to June 2011. There were 336 males and 229 females. They underwent gastroendoscopy examination due to upper gastrointestinal symptoms and had never received H. pylori eradication therapies. Duodenal ulcer patients were divided randomly into LACJ therapy group, LACB therapy group and LAC therapy group while gastritis patients LACJ therapy group and LACB therapy group. Group LAC received lansoprazole 30 mg + amoxicillin 1000 mg + clarithromycin 500 mg, twice a day, for 7 d (d1-7). Group LACJ: LAC therapy plus Jinghuaweikang, 3 capsules, twice a day, for 7 d (d1-7) then Jinghuaweikang, 3 capsules, twice a day, for 14 d (d8-21). Group LACB: LAC plus bismuth potassium citrate 220 mg, twice a day, for 7 d (d1-7) and then bismuth potassium citrate 220 mg, twice a day, for 14 d (d8-21). All duodenal ulcer patients received lansoprazole (30 mg, once a day) for 14 days after the first 7-day of treatment (d 8-21). At least 28 days after the end of treatment, all patients underwent (13)C urea breath test. Gastric mucosa was collected under endoscopy from the failed patients. The detection technique of gene chip was employed to detect antibiotics resistant gene from mucosa. RESULTS: The eradication rates of duodenal ulcer patients in groups LACJ, LACB and LAC were as follows: per-protocol (PP), 80.2% (77/96), 89.9% (89/99) and 72.2% (70/97) (P = 0.007), intention-to-treat (ITT), 78.6% (77/98), 88.1% (89/101) and 70.0% (70/100) (P = 0.007). No statistical differences existed between groups LACJ and LACB or LAC (all P > 0.05). But there were statistical differences between groups LACB and LAC (both P = 0.002). The eradication rates of PP and ITT of chronic gastritis patients in groups LACJ and LACB were as follows: 75.8% (97/128), 74.6% (97/130) vs 83.8% (109/130), 80.1% (109/136) (both P > 0.05). The symptomatic improvements of abdominal pain, burning and acid reflux of duodenal ulcer patients in group LACJ were higher than those in groups LACB and LAC. There were statistical differences between groups LACJ and LAC (all P < 0.05). The symptomatic improvements of bloating and belching for chronic gastritis patients in group LACJ were higher than those of group LACB. But no significant difference existed between two groups (all P > 0.05). Sixty samples of gastric mucosa were collected from the failed patients. The detection rates of antibiotic-resistant gene to clarithromycin and amoxicillin were 60.0% (36/36) and 18.3% (11/60) respectively. CONCLUSIONS: The efficacy of LACJ for the treatment of H. pylori infection patients is similar to LACB and superior to LAC. And the symptomatic improvement of patients is better than the other two regimens. The main cause of treatment failure is antibiotic resistance of H. pylori strains.

[Detection of APC gene germline mutation in Chinese familial adenomatous polyposis by direct sequencing in combination with multiplex ligation-dependent probe amplification].

OBJECTIVE: To investigate the characteristics of APC gene germline mutation in Chinese patients with familial adenomatous polyposis (FAP). METHODS: The genomic DNA was extracted from peripheral venous blood drawn from probands of 14 Chinese FAP families from Beijing, Hebei, Henan, Anhui, Inner Mongolia, Shanxi and Fujian. The APC gene was amplified by PCR and underwent direct sequencing. Large fragment deletion was detected by multiplex ligation-dependent probe amplification (MLPA) only in micromutation-negative samples found by sequencing. RESULTS: APC gene micromutations were found in 9 probands and the micromutation detection rate was 64.3%, including 6 frameshift mutations, 2 splicing mutations and 1 nonsense mutation. Large fragment deletions of APC gene were detected in 2 probands (14.3%). The total mutation detection rate of micromutation and large fragment deletion was 78.6%. Four novel micromutations and 2 novel large fragment deletions were found, including c.2336-2337insT, c.3923-3929delAAGAAAA, c.532-2A > T, c.4179-4180GAdelinsT, large fragment deletions of exons 11 and 10A and large fragment deletion of exon 15 start. CONCLUSIONS: In Chinese FAP patients, the germline mutation type of APC gene is variable; the majority is of frameshift mutation. The most common mutation site is exon 15. The mutation detection rate of APC gene can be boosted effectively by direct sequencing in combination with MLPA.

Molecular cloning of two genes encoding cinnamate 4-hydroxylase (C4H) from oilseed rape (Brassica napus).

Cinnamate 4-hydroxylase (C4H) is a key enzyme of phenylpropanoid pathway, which synthesizes numerous secondary metabolites to participate in development and adaption. Two C4H isoforms, the 2192-bp BnC4H-1 and 2108-bp BnC4H-2, were cloned from oilseed rape (Brassica napus). They both have two introns and a 1518-bp open reading frame encoding a 505-amino-acid polypeptide. BnC4H-1 is 57.73 kDa with an isoelectric point of 9.11, while 57.75 kDa and 9.13 for BnC4H-2. They share only 80.6% identities on nucleotide level but 96.6% identities and 98.4% positives on protein level. Showing highest homologies to Arabidopsis thaliana C4H, they possess a conserved p450 domain and all P450-featured motifs, and are identical to typical C4Hs at substrate-recognition sites and active site residues. They are most probably associated with endoplasmic reticulum by one or both of the N- and C-terminal transmembrane helices. Phosphorylation may be a necessary post-translational modification. Their secondary structures are dominated by alpha helices and random coils. Most helices locate in the central region, while extended strands mainly distribute before and after this region. Southern blot indicated about 9 or more C4H paralogs in B. napus. In hypocotyl, cotyledon, stem, flower, bud, young- and middle-stage seed, they are co-dominantly expressed. In root and old seed, BnC4H-2 is dominant over BnC4H-1, with a reverse trend in leaf and pericarp. Paralogous C4H numbers in Brassicaceae genomes and possible roles of conserved motifs in 5' UTR and the 2nd intron are discussed.