Results of Liquid Biopsy Studies by Next Generation Sequencing in Patients with Advanced Stage Non-small Cell Lung Cancer: Single Center Experience from Turkey.

Several studies demonstrated the utility of plasma-based cell-free circulating tumor DNA (ccfDNA) in determination of mutations in non-small cell lung cancer (NSCLC). We aimed to report our results of next generation sequencing (NGS) using liquid biopsy in patients with NSCLC. Patients with advanced stage NSCLC were enrolled and their genomic profiling results were recorded. Next generation sequencing targeted panel includes 19 hot-spot genes. The plasma was separated from the peripheral blood sample and ccfDNAs were isolated for NGS. We performed genomic profiling in 100 patients (20 females and 80 males) with a median age of 59.3 (range 26-79). A second liquid biopsy was performed in eight patients who developed progressive disease after the first treatment. The study population had adenocarcinoma (AC) (n = 73), squamous cell carcinoma (SCC) (n = 14), or NSCLC-NOS (not otherwise specified) (n = 13). In the SCC group, three of 14 patients had variants on EGFR and MET genes. In the AC and NSCLC-NOS groups, 39 out of 86 patients (45.3%) had variants. The most common one was in the EGFR gene (n = 27, 31.4%) including seven mutations related to drug resistance and two were polymorphisms. Three patients had both driver and resistance mutations (EGFR T790M, n = 2; KRAS exon 2 G12S and MET exon 14 E1012K, n = 1). Fifteen patients (17.4%) had an activating EGFR mutation and eight patients (9.3%) had variants in the KRAS gene. We reported our results regarding genomic profiling related to treatment using liquid biopsy in patients with NSCLC. Advantages of this method are the non invasiveness and reproducibility.

Factors associated with recurrent stroke and recanalization in patients presenting with isolated symptomatic carotid occlusion.

BACKGROUND AND PURPOSE: Patients with symptomatic internal carotid artery (ICA) occlusion constitute a small proportion of stroke/transient ischaemic attack patients who are at increased risk of early stroke recurrence and poor outcome. The optimal medical treatment for patients with symptomatic ICA occlusion who are ineligible for thrombolysis or thrombectomy is unknown. METHODS: Consecutive patients presenting at a single center with newly diagnosed symptomatic ICA occlusion (not involving the circle of Willis) were retrospectively reviewed. Those treated with intravenous thrombolysis or intra-arterial thrombolysis/thrombectomy were excluded. Patients were divided into two groups based on whether they experienced recurrent in-hospital stroke. RESULTS: The selected study population (n = 33) represented a small (20.4%) proportion of all newly symptomatic carotid occlusions, who nevertheless had an elevated risk of recurrent stroke during admission (24.2%). Of the variables examined (age, gender, admission National Institutes of Health Stroke Scale score, vascular risk factors, atrial fibrillation, prior stroke/transient ischaemic attack and anticoagulation within 48 h of presentation), only anticoagulation was significantly associated with a lower risk of in-hospital recurrent stroke. Anticoagulated patients showed a decreased incidence of stroke recurrence within the first week (6.7% vs. 38.9%, P = 0.032) and fewer strokes or deaths at 1 month (13.3% vs. 47.1%, P = 0.040). Hemorrhagic transformation was not observed in any patient. On follow-up imaging, ICA recanalization was significantly more frequent in anticoagulated patients (46.2% vs. 9.1%, P = 0.047). CONCLUSION: Patients with newly diagnosed symptomatic ICA occlusion (not involving the circle of Willis) represent a small but high risk subgroup of patients with carotid occlusion. Early anticoagulation was associated with fewer recurrent strokes and increased ICA recanalization. Larger scale prospective studies may be justified.

Evaluation of Health-related Quality of Life in Paediatric Patients with Acute Lymphoblastic Leukaemia.

OBJECTIVE: Health-related quality of life (HRQOL) measurement is used for assessing the impact of diseases and medical treatments on physical, psychological and social aspects of an individual's health and life. The Paediatric Quality of Life Inventory™ (PedsQL™) is a widely used instrument to measure paediatric HRQOL in children. The aim of this study is to investigate the HRQOL in paediatric patients with acute lymphoblastic leukaemia and determine the precautions for improving the quality of their life. METHODS: Paediatric Quality of Life Inventory™ 4.0 was administered to 75 paediatric patients with acute lymphoblastic leukaemia, and 50 healthy age- and gender-matched children. RESULTS: Health-related quality of life scores were significantly lower in patients with acute lymphoblastic leukaemia than in healthy controls in this study. CONCLUSIONS: It is thought that determination of the psychosocial, as well as the physical impacts of the disease on the child, will positively influence the treatment given by improving the quality of life of both the child and the family.

New method for estimating the volume and volume fractions of the nasal structures in the goose (Anser anser domesticus) using computed tomography images.

1. The conchae within the nasal cavity of poultry are important for water and energy conservation, but have not been experimentally evaluated. The aim of the present study was to determine the accuracy of volume and volume fraction estimates of the conchae, nasal septum and nasal cavity. 2. The nasal cavities of 7 adult goose heads were scanned using computed tomography (CT), with images sampled randomly at a 1/5 sampling fraction. Physical sections were obtained from the same samples, using an electric saw that had an adjustable section range, and provided 14 to 15 sections with a thickness of 2.5 mm. The section surface areas of the nasal cavity, nasal septum and conchae were estimated using the Cavalieri principle. Results obtained using the CT and physical section images were compared. Volumes and volume fractions obtained from the physical sections were accepted as the gold standard and differences in the CT images were determined. 3. Multiplication of the data obtained on the CT images with the deviation percentage of the physical sections produced normalised values. No differences were observed between the gold standard data and the CT images. While it was possible to normalise the obtained data using the gold standard values, the raw data could also be used for comparative studies because the deviations from normal would be similar for all groups. 4. Our study showed that the nasal structures could be estimated in vivo using CT images.

PTEN, Akt, MAPK, p53 and p95 expression to predict trastuzumab resistance in HER2 positive breast cancer.

PURPOSE: Mutations that activate the PIK3CA oncogene and inhibit the tumor suppressor gene PTEN action are commonly found in breast tumors. Akt is a key activator of cell survival. p53 is frequently found mutated in human tumors, and mutant p53 protein actively contributes to tumorigenesis. In selected cases of breast cancer, trastuzumab (TZMB) is incorporated in the primary treatment in the adjuvant and metastatic settings. Many studies have reported that selected patients are resistant to TZMB due to the presence of p95 HER2 fragments. To address this, we analysed PTEN, Akt, MAPK, p53 and p95 expression in breast cancer patients treated with TZMB. METHODS: Out of 90 patients histologically diagnosed with breast cancer between 2004 and 2011, analysed were 25 patients with HER2 positive, and estrogen (ER) and progesterone receptors (PR) negative, metastatic or locally advanced disease. All 25 patients were treated with TZMB and resistance to TZMB was assessed. All patients were on anthracycline-and taxane-containing regimens. Tissue samples were obtained from paraffin blocks and evaluated immunohistochemically for PTEN, Akt, MAPK, p53, and p95 expression. RESULTS: TZMB resistance was detected in 5 (20%) patients. Akt expression was positive in 2 patients (8%) and MAPK, p95, and p53 expression was positive in 1 patient (4%); PTEN expression was negative in 3 patients (12%). No significant differences were found between TZMB resistance and PTEN, Akt, MAPK, p53, and p95 expression. Subgroup analysis was carried out in the neoadjuvant treatment group. Complete pathologic response was detected in 3 patients (21.4%). Statistically significant differences were not found between the complete response rate and PTEN, Akt, MAPK, and p95 expression. There was a statistically significant correlation between p53 expression and complete pathologic response (p=0.02). CONCLUSION: No statistically significant correlation between TZMB resistance and the expression of these biomarkers was noted. In patients with HER2-positive breast cancer that were treated with 4 dose-dense sequential cycles of doxorubicin and cyclophosphamide, followed by TZMB and paclitaxel combination therapy in the neodjuvant setting, p53 expression could predict complete response to chemotherapy.

Awareness and obscurity: demystifying genomic profiling in real-world practice for the members of the Turkish Society of Medical Oncology (TSMO).

OBJECTIVE: Molecular testing in oncology practice is increasingly being used to offer more relevant therapies to cancer patients. Our study aims to determine the real-world impact of routine incorporation of molecular testing among the Turkish Oncology community across all types of cancer and identify gaps for the first time. SUBJECTS AND METHODS: This research was conducted in Turkey among medical oncologists from different backgrounds. The survey attendance was entirely voluntary. A questionnaire with twelve items (multiple choice +/- closed-ended) was utilized in this study to assess the effect of molecular tests in real clinical situations. RESULTS: 102 oncologists with various levels of experience participated in this study. Most of the respondents (97%) reported successful implementation of molecular testing. About 10% of the participating oncologists said they preferred genetic tests at the early stages of cancer, compared to the majority who preferred genetic tests at the terminal stage. Molecular tests are often performed in separate locations and 47% of the oncologists were using a targeted panel specific to the type of malignancy. CONCLUSIONS: Several informational difficulties must be resolved in order to have early personalized therapy as the standard treatment. We need accessible, comprehensive, and regularly updated databases to compare genetic profiling and its therapeutic implications. We also need to continue educating patients and physicians.

Estimation of nasal cavity and conchae volumes by stereological method.

BACKGROUND: Studies evaluating the mean volumes of nasal cavity and concha are very rare. Since there is little date on the mentioned topic, we aimed to carry out the presented study to obtain a volumetric index showing the relation between the nasal cavity and concha. MATERIAL AND METHODS: The volumes of the nasal cavity and concha were measured in 30 males and 30 females (18-40 years old) on computed tomography images using stereological methods. RESULTS: The mean volumes of nasal cavity, concha nasalis media, and concha nasalis inferior were 5.95 ± 0.10 cm(3), 0.56 ± 0.22 cm(3), and 1.45 ± 0.68 cm(3); 7.01 ± 0.18 cm(3), 0.67 ± 0.31 cm(3) and 1.59 ± 0.98 cm(3) in females and males, respectively. There were statistically significant differences in the volume of the nasal cavity and concha nasalis media (p 〈 0.05) between males and females, except for concha nasalis inferior (p 〉 0.05). CONCLUSIONS: Our results could provide volumetric indexes for the nasal cavity and concha, which could help the physician to manage surgical procedures related to the nasal cavity and concha.

Nurse-assisted education and exercise decrease the prevalence and morbidity of lymphedema following breast cancer surgery.

PURPOSE: To evaluate an educational and exercise program for the prevention and progression of post-mastectomy lymphedema of the arm and shoulder. METHODS: Fifty-five patients who had undergone mastectomy and axillary lymph node dissection between June 2009 and January 2010 were included in this study. The patients were informed by a trainer nurse about the precautions they should take to prevent the development of lymphedema. The patients were also trained for the appropriate exercises and were given written educational material prepared by the investigators. RESULTS: Among the participants, 96.4% underwent modified radical mastectomy (MRM) and 3.6% breast-conserving (BCS) surgery. The mean postoperative follow-up period was 9.87 ± 17.55 months. The degree of lymphedema was found lower, even within 6 months, in the patients that exercised as compared to the patients that did not (p<0.05). CONCLUSIONS: The results indicate that the risk of development and progression of mastectomy-related lymphedema was reduced with education and exercise provided by trained nurses at an early stage.

Patterns of COVID-19-related headache: A cross-sectional study.

BACKGROUND: Headache is the most common COVID-19-related neurological symptom. We investigated the characteristics of COVID-19-related headache and their relationship with clinical severity in Kirsehir Province, Turkey. METHODS: This cross-sectional study prospectively enrolled 226 COVID-19-positive patients who developed headache during acute infection. Demographic data, headache characteristics, and infection symptoms were recorded. The clinical severity of COVID-19 was documented in each participant. RESULT: New-onset COVID-19-related headaches lasting 4 days were reported in 164 patients (72.5 %); these were mostly bilaterally or localized to the forehead (58.4 %), pulsating (42.5 %), moderate to severe intensity (30.1 %), with a partial response to paracetamol (23.5 %). The other 62 patients (27.4 %) reported headaches before COVID-19. Their COVID-related headaches were fiery type (p = 0.025), of very severe intensity (p = 0.008), had a holocranial distribution (p = 0.004), and were less response to paracetamol (p = 0.003); the headaches were significantly more frequent after COVID-19 than before COVID-19. Older age, high body mass index, and low education level were significantly higher in the severe group (all p < 0.001). Female sex (p = 0.019) and being a healthcare worker (p < 0.001) were significantly more frequent in mild cases. CONCLUSIONS: Bilateral, prolonged, moderate to severe headaches that were analgesic resistant are more frequent in patients with COVID-19 infection. Further study should examine whether the headache characteristics distinguish COVID-19-related headaches from other types, particularly in asymptomatic subjects.

Plasma biomarkers of brain injury in COVID-19 patients with neurological symptoms.

OBJECTIVE: Neurological symptoms (NS) were often reported in COVID-19 infection. We examined the plasma levels of glial fibrillary acidic protein (GFAP) and S100B together, as brain injury biomarkers, in relation to persistent NS in a cohort of patients with COVID-19 during the acute phase of the disease. METHODS: A total of 20 healthy controls and 58 patients with confirmed COVID-19 were enrolled in this prospective study. Serum GFAP and S100B levels were measured by using enzymle linked immunoassay method from blood samples. RESULTS: Serum GFAP levels were found to be significantly higher in the severe group than in the controls (p = 0.007). However, serum S100B levels were similar between control and disease groups (p > 0.05). No significant results for GFAP and S100B were obtained between the disease groups depending on whether the sampling time was below or above 5 days (p > 0.05). We did not find a correlation between serum GFAP and S100B levels and the presence of NS (p > 0.05). However, serum S100B levels were slightly higher in patients with multiple NS than in those with a single symptom (p = 0.044). CONCLUSIONS: Elevated GFAP was associated with disease severity but not with NS in COVID-19 patients. Whereas, high serum S100B was associated with the multipl NS in these patients. Our data suggest that GFAP and S100B may be of limited value currently in order to represent the neuronal damage, though serving a basis for the future work.

Endoscopic treatment of biliary disorders in patients with Roux-en-Y hepaticojejunostomy via a permanent access loop.

The management of biliary disorders in patients with Roux-en-Y hepaticojejunostomy anastomosis is challenging and remains controversial. Our aim is to share our experiences of endoscopic treatment via a permanent access loop in 5 patients. Endoscopic treatment via a permanent access loop is an invaluable procedure for the management of stenotic hepaticojejunostomy anastomosis, anastomotic leakage, and hepatolithiasis. It may even be life-saving for some patients.

Nasal secretory protein changes following intravenous choline administration in calves with experimentally induced endotoxaemia.

Nasal secretory fluid proteomes (NSPs) can provide valuable information about the physiopathology and prognosis of respiratory tract diseases. This study aimed to determine changes in NSP by using proteomics in calves treated with lipopolysaccharide (LPS) or LPS + choline. Healthy calves (n = 10) were treated with LPS (2 µg/kg/iv). Five minutes after LPS injection, the calves received a second iv injection with saline (n = 5, LPS + saline group) or saline containing 1 mg/kg choline (n = 5, LPS + choline group). Nasal secretions were collected before (baseline), at 1 h and 24 h after the treatments and analysed using label-free liquid chromatography-tandem mass spectrometry (LCMS/MS). Differentially expressed proteins (>1.2-fold-change) were identified at the different time points in each group. A total of 52 proteins were up- and 46 were downregulated at 1 h and 24 h in the LPS + saline group. The upregulated proteins that showed the highest changes after LPS administration were small ubiquitin-related modifier-3 (SUMO3) and glutathione peroxidase-1 (GPX1), whereas the most downregulated protein was E3 ubiquitin-protein ligase (TRIM17). Treatment with choline reduced the number of upregulated (32 proteins) and downregulated proteins (33 proteins) in the NSPs induced by LPS. It can be concluded that the proteome composition of nasal fluid in calves changes after LPS, reflecting different pathways, such as the activation of the immunological response, oxidative stress, ubiquitin pathway, and SUMOylation. Choline treatment alters the NSP response to LPS.

Drainless resection of the submandibular gland with facial vessel preservation: A comparative study.

OBJECTIVE: Resection of the submandibular gland (SMG) is a common surgery and many surgeons leave a drainage system in the surgical field. However, surgical drain may increase risk of complication and related prolonged hospitalization time. The purpose of this study was to investigate the safety of SMG resection without any surgical drainage system. METHODS: This retrospective trial on SMG surgery was conducted between 2016 and 2019. Patients were assigned into one of two main groups: surgical drain (+) (n=20) and surgical drain (-) (n=17). All surgical procedures were done via a standardized surgical technique. Facial vessels were dissected and only glandular branches were ligated. Also, non-identification method was applied for marginal mandibular nerve (MMN) protecting. In surgical drain (-) group, before the wound closure, oxidized regenerated cellulose (ORC) was placed in the surgical field. Moreover, a closed suction drain was inserted in surgical drain (+) group. RESULTS: A total of 37 SMG resections were performed: 15 patients had sialolithiasis, 14 patients had pleomorphic adenoma and 8 patients had chronic sialadenitis. There were 20 women (54%) and 17 men (46%), with an age range of 23-70 years. No major complications were observed in surgical drain (-) group. There were two cases with minor complications. One patient (5.8%) occurred transient paralysis of the MMN. Other patient developed seroma and it was easily managed with repeated punctures. ORC related allergic reaction or adverse incident were not detected in any of the patients. On the other hand, in surgical drain (+) group, 2 patients (10%) developed a hematoma on the first postoperative day and local wound infection was detected in 4 patients (20%). We found that the surgical drain usage was related to prolonged hospitalization, worse wound healing and problems with scarring. CONCLUSION: Our findings provide evidence for the safe drainless resection of the SMG using ORC. It may be possible to prevent all of these undesirable conditions by a surgery which performed without drain insertion.

Mucormycosis-associated fungal infections in patients with haematologic malignancies.

Among patients with haematologic disorders, mucormycosis most commonly occurs in those with acute leukaemia or lymphoma who have developed neutropenia due to malignancy or to chemotherapy, and in transplanted patients receiving immunosuppressive treatment. Here, we aim to present a retrospective study conducted over a 5-year period (2001-2005). The study included 20 patients with haematologic malignancies with a proven mucormycosis admitted in Medical Oncology Divisions in Cukurova University Hospital. The most frequent sites of infection were paranasal sinuses (95%) and lung (5%). Antifungal treatment was empirically administered in 18 (90%) patients; 18 patients underwent radical surgical debridement (90%). The therapy was successful for only eight patients (40%). Eleven patients died within 1 months of the diagnosis of fungal infection: the cause of death was only by mucormycosis in four patients (36.6%), mucormucosis and systematic inflamatuar response syndrome (SIRS) in two patients (18.2%) and progression of haematologic disease in five patients (45.5%). At univariate analysis, the factors that correlated with a positive outcome from infection were the following: amphotericin B treatment, neutrophil recovery from postchemotherapy aplasia. At multivariate analysis, the factors that significantly correlated with recovery from infection were the liposomal amphotericin B treatment (p = 0.026), doses of L-AmB (p = 0.008) and the length of the treatment (p = 0.01), respectively. It seems to have increased in recent years. Although a reduction of mortality has been observed recently, the mortality rate still remains high. Extensive and aggressive diagnostic and therapeutic procedures are essential to improve the prognosis in these patients.

On demand pancreatic stenting in chronic pancreatitis might provide good palliation of pain.

BACKGROUND AND AIM: Chronic pancreatitis (CP)-related pain is a considerable problem in gastroenterology practice that frequently requires several endoscopic interventions. We aimed to investigate the efficacy of pancreatic duct stenting performed on demand, instead of at defined intervals, for the management of the CP-related pain. METHODS: This study is a retrospective evaluation of thirteen years of data. Sixty-seven patients with CP who suffered from intractable pain were enrolled in the study. Pancreatic stenting was performed mainly with single stents according to the diameter of the pancreatic duct and width of the stricture or, less frequently, with multiple stents aiming to achieve stricture resolution. The subsequent endoscopic session was scheduled based on the patient's symptoms. RESULTS: Overall, 65 of 67 patients underwent successful pancreatic cannulation (technical success rate 97%). Fifty-seven patients with a pancreatic stenting history were still undergoing follow-up. Of these patients, 26 patients still had pancreatic ductal stents; however, the stents were removed from 31 patients. Only 8 patients (25%) required further endoscopic or surgical intervention because of the re-emergence of pain after a median stent-free period of 17 months (3-127 months). One patient with a biliary stricture and one patient with a pancreatic mass underwent surgery. Pancreatic stents remained for a median length of 14 months (3-84 months). During the follow-up period, 55 of 65 patients became pain-free or had partial pain relief (clinical success rate 84%). CONCLUSIONS: On demand replacement of pancreatic stent is feasible in patients with CP and it might provide a good palliation of CP-related pain.

An efficient stereological sampling approach for quantitative assessment of nerve regeneration.

AIMS: The aim of the present study was to find the most efficient sampling strategy for stereological analysis of peripheral nerve, including the number of myelinated nerve fibres, axon cross-sectional area and myelin sheet thicknesses of nerve fibres. METHODS: Two groups of rats underwent experimental resection of the tibial and peroneal nerves. The first group received tibial-peroneal end to end autograft repair (n = 6). Tibial and peroneal nerves were isolated, transected, and separated 1 cm distal to the trifurcation, where they lay adjacent to each other by a 1-cm gap, then repaired with an autologous nerve graft taken from the tibial nerve. The proximal stump of the tibial nerve and distal stump of the peroneal nerve were connected to each other by means of the nerve graft. The second group received tibial-peroneal repair with a flexible collagen tube (n = 6). After 90 days of recovery, animals were sacrificed and nerve segments were removed and sectioned for microscopy. Three different sampling strategies, that is, small, medium and large step sizes were applied to obtain each quantitative parameter. RESULTS: There are no significant differences between these sampling strategies with respect to total number of myelinated nerve fibres, axon cross-sectional area and myelin sheet thicknesses of nerve fibres. CONCLUSION: Findings show that one can achieve the desired estimate precisely with a rather large and less time-consuming sampling approach. In addition, it was observed that the size discrepancy of nerve regeneration can be improved by collagen tube conduit even with a 1-cm gap.

The fastest animals and vehicles are neither the biggest nor the fastest over lifetime.

Here we show how the size of a body affects its maximum average speed of movement through its environment. The theoretical challenge was to predict that 'outliers' must exist, such as the cheetah for terrestrial animals and the jet fighter for airplanes. We show that during a travel that starts from rest and continues at cruising speed, the body size for minimum travel time, or maximum average speed, is not the biggest. The results are compared with extensive data for military aircraft for chase, attack and reconnaissance, in addition to data for commercial aircraft. The paper also explains why in earlier studies of flying (animals, airplanes) the airplane data deviated upward (toward greater speeds) relative to the theoretical trend followed by flying animals, and why the fastest animal flyers are one thousand times smaller than the fastest swimmers. Unlike the biggest animals and airplanes (elephant, whale, commercial jet), which move constantly, the fastest animals and airplanes spend most of their lives at rest. When judged for speed averaged over lifetime, the fastest 'sprinters' are in fact the slowest movers (as in Aesop's fable 'The Tortoise and the Hare').

Endoscopic treatment of pancreatic fistulas.

BACKGROUND: Pancreatic fistulas are managed primarily by conservative treatment. Surgery is performed in cases of conservative treatment failure. Endoscopic treatment is reported to be both effective and safe as an alternative treatment method. METHODS: A total of 26 patients underwent endoscopic treatment after failure of conservative treatment between January 2002 and November 2004. The mean time between the onset of fistula and the endoscopic retrograde cholangiopancreatography (ERCP) procedure was 95 days. The mean fistula output volume was 400 ml per day. Four patients had pancreatic ascites. The aim of the endoscopic treatment was to bypass the ductal disruption by placing stents or drains where the origin of fistulous tract could be identified, and to lower the pancreatic duct pressure by performing pancreatic sphincterotomy or by placing stents where the site of the leak could not be identified. RESULTS: Pancreatography could be performed in all the patients except one. Partial duct disruption occurred in 16 patients. All of the fistulas closed after the ductal disruption was bypassed. Pancreatic sphincterotomy or endoprothesis placement was effective for eight of the remaining nine patients in whom the ductal disruption originated from the tail of the pancreas and hence could not be bypassed. The overall success rate was 94% for the patients with partial duct disruption. Four patients had side branch leaks. All of them closed after placement of an endoprothesis. Fistulas closed in only one (20%) of the five patients with complete duct disruption. Pancreatic ascites resolved in two of the four patients after endoscopic treatment. No serious complications resulted from endoscopic treatment other than proximally migrated stents in two patients. CONCLUSIONS: Endoscopic treatment is an effective and safe method for patients with pancreatic fistulas unresponsive to conservative treatment. The success rate is very high, especially for patients with partial and side branch duct disruption.