Lumbar lordosis restoration by minimally invasive short-segment fusion with anterior column realignment for adult spinal deformity: minimum 2-year follow-up.

OBJECTIVE: The efficacy of anterior column realignment (ACR) remains relatively unclear, possibly because some safety concerns have limited its adoption and extensive evaluation. The authors aimed to study whether a minimally invasive surgery (MIS) triad consisting of ACR, lateral lumbar interbody fusion, and percutaneous pedicle screw fixation in a select group of adult spinal deformity (ASD) patients helps shorten fusion length without compromising clinical and radiographic outcomes over a minimum 2-year follow-up period. METHODS: A series of 61 ASD patients (mean age 72.8 years) with pelvic incidence (PI) - lumbar lordosis (LL) (PI-LL) mismatch > 10° underwent the short-segment MIS triad (mean fusion length 3.0 levels) as a single-stage operation with a mean operative time and estimated blood loss of 157 minutes and 127 mL, respectively. Exclusion criteria were 1) thoracic scoliosis as the main deformity, 2) thoracolumbar junction kyphosis > 25°, 3) ankylosed facet joints, and 4) previous spinal fusion surgery. Seven patients, who needed fusion to be extended to S1, underwent mini-open transforaminal lumbar interbody fusion at L5-S1. RESULTS: The segmental disc angle at the ACR level more than quintupled, averaging from 2.9° preoperatively to 18.9° at the latest follow-up (p < 0.0001). LL, in turn, nearly doubled from 17.0° to 32.8° (p < 0.0001) and PI-LL decreased by nearly half from 28.8° to 13.2° (p < 0.0001). At the same time, other spinopelvic deformity parameters as well as Oswestry Disability Index (ODI) scores significantly improved. Patients were divided into two groups at the latest postoperative evaluation: 36 patients whose PI-LL improved to < 10° and 25 patients who maintained a PI-LL mismatch > 10°. Binary logistic regression revealed preoperative PI-LL mismatch as the only factor that significantly influenced this dichotomous separation postoperatively. Receiver operating characteristic curve analysis identified the critical preoperative mismatch of 26.4° with 68% sensitivity and 84% specificity. Despite this different radiographic consequence, the two groups had an equally successful clinical outcome with no significant difference in ODI scores. CONCLUSIONS: As long as the ASD characteristics are consistent with the authors' exclusion criteria, the short-segment MIS triad served as an excellent surgical option in the patients with preoperative PI-LL mismatch < 26.4°, but the technique also worked well even in those with a mismatch > 26.4°, although ideal spinopelvic alignment targets were not necessarily achieved in these patients.

New Effective Intraoperative Techniques for the Prevention of Coronal Imbalance after Circumferential Minimally Invasive Correction Surgery for Adult Spinal Deformity.

This study aimed to devise measures and investigate their effect on coronal imbalance (CI) after circumferential minimally invasive correction surgery (CMIS) with lateral lumbar interbody fusion and percutaneous pedicle screw for adult spinal deformity (ASD). A total of 115 patients with ASD who underwent CMIS from the lower thoracic spine to the ilium were included. Patients were stratified based on the distance between the spinous process of the upper instrumented vertebra and central sacrum vertical line (UIV-CSVL) after the first intraoperative rod application into groups P (UIV-CSVL > 10 mm, n = 50) and G (UIV-CSVL < 10 mm, n = 65). Measures to correct postoperative CI introduced during surgery, preoperative and postoperative UIV-CSVL, and changes in UIV-CSVL after various measures (ΔUIV-CSVL) were investigated in group P. Rod rotation (RR), S2 alar-iliac screw distraction (SD), and kickstand-rod (KR) technique were performed in group P. Group P was further divided into group RR (n = 38), group SD (RR and SD) (n = 7), and group KR (RR and KR) (n = 5); the ΔUIV-CSVLs were 13.9 mm, 20.1 mm, and 24.4 mm in these three groups, respectively. Postoperative C7-CSVL < 10 mm was achieved in all three correction groups. In conclusion, our measures enabled sufficient correction of the UIV-CSVL and are useful for preventing CI after CMIS for ASD.

Surgical Approaches and Short-Term Results of Circumferential Minimally Invasive Correction Surgery for Adult Idiopathic Scoliosis.

OBJECTIVE: We present the surgical approaches and short-term (2 years postoperative) results pertaining to circumferential minimally invasive spine surgery (CMIS) with lateral lumbar interbody fusion (LLIF) and percutaneous pedicle screw application for adult idiopathic scoliosis (AS). METHODS: We enrolled eight patients with AS who underwent CMIS (2018-2020) and examined the number of fused levels, upper instrumented vertebra, lower instrumented vertebra, number of LLIF-treated segments, number of preoperative intervertebral fusions, intraoperative blood loss, operative time, various spinopelvic parameters, Oswestry Disability Index, low back pain, visual analog scale (VAS), leg VAS, bone fusion rate, and perioperative complications. RESULTS: The upper instrumented vertebra was T4, T7, T8, and T9 in two cases, whereas lower instrumented vertebra was the pelvis in all the cases. The average numbers of fixed vertebrae and segments that underwent LLIF were 13.3 ± 2.0 and 4.6 ± 0.7, respectively. All spinopelvic parameters improved significantly after surgery (thoracic kyphosis: P < 0.05, lumbar lordosis, cobb angle, pelvic tilt, pelvic incidence-lumbar lordosis, sagittal vertical axis: P < 0.001), and good alignment was achieved. The Oswestry Disability Index and VAS scores improved significantly (P < 0.001). The bone fusion rates achieved in the lumbosacral and thoracic spine were 100% and 88%, respectively. Only 1 patient showed postoperative coronal imbalance. CONCLUSIONS: The 2-year postoperative results of CMIS for AS were good, and spontaneous bone fusion was confirmed in the thoracic spine without bone grafting. In this procedure, sufficient intervertebral release with LLIF and a percutaneous pedicle screw device translation technique enabled adequate global alignment correction. Therefore, correcting the global imbalance of the coronal and sagittal planes is more crucial than correcting scoliosis.

Visualization of axonal and volume currents in median nerve compound action potential using magnetoneurography.

OBJECTIVE: Reconstruct compound median nerve action currents using magnetoneurography to clarify the physiological characteristics of axonal and volume currents and their relationship to potentials. METHODS: The median nerves of both upper arms of five healthy individuals were investigated. The propagating magnetic field of the action potential was recorded using magnetoneurography, reconstructed into a current, and analyzed. The currents were compared with the potentials recorded from multipolar surface electrodes. RESULTS: Reconstructed currents could be clearly visualized. Axonal currents flowed forward or backward in the axon, arcing away from the depolarization zone, turning about the subcutaneous volume conductor, and returning to the depolarization zone. The zero-crossing latency of the axonal current was approximately the same as the peak of its volume current and the negative peak of the surface electrode potential. Volume current waveforms were proportional to the derivative of axonal ones. CONCLUSIONS: Magnetoneurography allows the visualization and quantitative evaluation of action currents. The currents in axons and in volume conductors could be clearly discriminated with good quality. Their properties were consistent with previous neurophysiological findings. SIGNIFICANCE: Magnetoneurography could be a novel tool for elucidating nerve physiology and pathophysiology.

Magnetoneurography to investigate the mechanisms underlying the P9 far-field potential.

OBJECTIVE: The mechanism underlying the generation of P9 far-field somatosensory evoked potentials (SEPs) is unresolved. Accordingly, we used magnetoneurography to visualize the current distribution in the body at the P9 peak latency and elucidate the origin of P9 generation. METHODS: We studied five healthy male volunteers without neurological abnormalities. We recorded far-field SEPs after median nerve stimulation at the wrist to identify the P9 peak latency. Using magnetoneurography, we recorded the evoked magnetic fields in the whole body under the same stimulus conditions as the SEP recording. We analyzed the reconstructed current distribution at the P9 peak latency. RESULTS: At the P9 peak latency, we observed the reconstructed current distribution dividing the thorax into two parts, upper and lower. Anatomically, the depolarization site at the P9 peak latency was distal to the interclavicular space and at the level of the second intercostal space. CONCLUSIONS: By visualizing the current distribution, we proved that P9 peak latency originates in the change in volume conductor size between the upper and lower thorax. SIGNIFICANCE: We clarified that magnetoneurography analysis is affected by the current distribution due to the junction potential.

A triple minimally invasive surgery combination for subacute osteoporotic lower lumbar vertebral collapse with neurological compromise: a potential alternative to the vertebral corpectomy/expandable cage strategy.

OBJECTIVE: Acute/subacute osteoporotic vertebral collapses (OVCs) in the lower lumbar spine with neurological compromise, although far less well documented than those in the thoracolumbar junction, may often pose greater treatment challenges. The authors clarified the utility of 3 familiar combined techniques of minimally invasive surgery for this condition as an alternative to the corpectomy/expandable cage strategy. METHODS: This report included the authors' first 5 patients with more than 2 years (range 27-48 months) of follow-up. The patients were between 68 and 91 years of age, and had subacute painful L4 OVC with neurological compromise and preexisting lumbar spinal stenosis. The authors' single-stage minimally invasive surgery combination consisted of the following: step 1, balloon kyphoplasty for the L4 OVC to restore its strength, followed by L4-percutaneous pedicle screw (PPS) placement with patients in the prone position; step 2, tubular lateral lumbar interbody fusion (LLIF) at the adjacent disc space involved with endplate injury, with patients in the lateral position; and step 3, supplemental PPS-rod fixation with patients in the prone position. RESULTS: Estimated blood loss ranged from 20 to 72 mL. Neither balloon kyphoplasty-related nor LLIF-related potentially serious complications occurred. With CT measurements at the 9 LLIF levels, the postoperative increases averaged 3.5 mm in disc height and 3.7 mm in bilateral foraminal heights, which decreased by only 0.2 mm and 0 mm at the latest evaluation despite their low bone mineral densities, with a T-score of -3.8 to -2.6 SD. Canal compromise by fracture retropulsion decreased from 33% to 23% on average. As indicated by MRI measurements, the dural sac progressively enlarged and the ligamentum flavum increasingly shrank over time postoperatively, consistent with functional improvements assessed by the physician-based, patient-centered measures. CONCLUSIONS: The advantages of this method over the corpectomy/expandable cage strategy include the following: 1) better anterior column stability with a segmentally placed cage, which reduces stress concentration at the cage footplate-endplate interface as an important benefit for patients with low bone mineral density; 2) indirect decompression through ligamentotaxis caused by whole-segment spine lengthening with LLIF, pushing back both the retropulsed fragments and the disc bulge anteriorly and unbuckling the ligamentum flavum to diminish its volume posteriorly; and 3) eliminating the need for segmental vessel management and easily bleeding direct decompressions. The authors' recent procedural modification eliminated step 3 by performing loose PPS-rod connections in step 1 and their tight locking after LLIF in step 2, reducing to only once the number of times the patient was repositioned.

Radiographic and MRI evidence of indirect neural decompression after the anterior column realignment procedure for adult spinal deformity.

OBJECTIVE: The anterior column realignment (ACR) procedure, which consists of sectioning the anterior longitudinal ligament/annulus and placing a hyperlordotic interbody cage, has emerged as a minimally invasive surgery (MIS) for achieving aggressive segmental lordosis enhancement to address adult spinal deformity (ASD). Although accumulated evidence has revealed indirect neural decompression after lateral lumbar interbody fusion (LLIF), whether ACR serves equally well for neural decompression remains to be proven. The current study intended to clarify this ambiguous issue. METHODS: A series of 36 ASD patients with spinopelvic mismatch, defined as pelvic incidence (PI) minus lumbar lordosis (LL) > 10°, underwent a combination of ACR, LLIF, and percutaneous pedicle screw (PPS) fixation. This "MIS triad" procedure was applied over short segments with mean fusion length of 3.3 levels, and most patients underwent single-level ACR. The authors analyzed full-length standing radiographs, CT and MRI scans, and Oswestry Disability Index (ODI) scores in patients with minimum 1 year of follow-up (mean [range] 20.3 [12-39] months). RESULTS: Compared with the preoperative values, the radiographic and MRI measurements of the latest postoperative studies changed as follows. Segmental disc angle more than quadrupled at the ACR level and LL nearly doubled. MRI examinations at the ACR level revealed a significant (p < 0.0001) increase in the area of the dural sac that was accompanied by significant (p < 0.0001) decreases in area and thickness of the ligamentum flavum and in thickness of the disc bulge. The corresponding CT scans demonstrated significant (all p < 0.0001) increases in disc height to 280% of the preoperative value at the anterior edge, 224% at the middle edge, and 209% at the posterior edge, as well as in pedicle-to-pedicle distance to 122%. Mean ODI significantly (p < 0.0001) decreased from 46.3 to 26.0. CONCLUSIONS: The CT-based data showing vertebral column lengthening across the entire ACR segment with an increasingly greater degree anteriorly suggest that the corrective action of ACR relies on a lever mechanism, with the intact facet joints acting as the fulcrum. Whole-segment spine lengthening at the ACR level reduced the disc bulge anteriorly and the ligamentum flavum posteriorly, with eventual enlargement of the dural sac. ACR plays an important role in not only LL restoration but also stenotic spinal canal enlargement for ASD surgery.

A New Treatment Algorithm That Incorporates Minimally Invasive Surgery for Pyogenic Spondylodiscitis in the Thoracic and Lumbar Spines: The Results of Its Clinical Application to a Series of 34 Patients.

Background and Objectives: Spinal minimally invasive surgery (MIS) experts at the university hospital worked as a team to develop a new treatment algorithm for pyogenic spondylodiscitis in lumbar and thoracic spines. They modified a flow chart introduced for this condition in a pre-MIS era to incorporate MIS techniques based on their extensive experiences accumulated over the years, both in MIS for degenerative lumbar diseases and in the treatment of spine infections. The MIS procedures incorporated in this algorithm consisted of percutaneous pedicle screw (PPS)-rod fixation and transpsoas lateral lumbar interbody fusion (LLIF). The current study analyzed a series of 34 patients treated with prospective selection of the methods according to this new algorithm. Materials and Methods: The algorithm first divided the patients into those who had escaped complicated disease conditions, such as neurologic impairment, extensive bone destruction, and the need to be mobilized without delay (Group 1) (19), and those with complicated pyogenic spondylodiscitis (Group 2) (15). Group 1 had image-guided needle biopsy followed by conservative treatment alone with antibiotics and a spinal brace (12) (Group 1-A) or a subsequent addition of non-fused PPS-rod fixation (7) (Group 1-B). Group 2 underwent an immediate single-stage MIS with non-fused PPS-rod fixation followed by posterior exposure for decompression and debridement through a small midline incision (12) (Group 2-A) or an additional LLIF procedure after an interval of 3 weeks (3) (Group 2-B). Results: All patients, except four, who either died from causes unrelated to the spondylodiscitis (2) or became lost to follow up (2), were cured of infection with normalized CRP at an average follow up of 606 days (105-1522 days). A solid interbody fusion occurred at the affected vertebrae in 15 patients (50%). Of the patients in Group 2, all but two regained a nearly normal function. Despite concerns about non-fused PPS-rod instrumentation, only seven patients (21%) required implant removal or replacement. Conclusions: Non-fused PPS-rod placements into infection-free vertebrae alone or in combination with posterior debridement through a small incision worked effectively in providing local stabilization without contamination of the metal implant from the infected tissue. MIS LLIF allowed for direct access to the infected focus for bone grafting in cases of extensive vertebral body destruction.

Bone Fusion Morphology after Circumferential Minimally Invasive Spine Surgery Using Lateral Lumbar Interbody Fusion and Percutaneous Pedicle Screws without Bone Grafting in the Thoracic Spine: A Retrospective Study.

Background and Objectives: This study aimed to investigate the process and morphology of thoracic and lumbosacral bone fusion in patients with adult spinal deformity (ASD) who underwent circumferential minimally invasive spine surgery (CMIS) by lateral lumbar interbody fusion (LLIF) and percutaneous pedicle screws (PPSs) without bone grafting in the thoracic spine and who have risk factors for bone fusion failure in the thoracic spine. Materials and Methods: This retrospective study included 61 patients with spinal deformities (46 women and 15 men) who underwent CMIS with LLIF and PPSs at our hospital after 2016 and completed a 3-year postoperative follow-up. The rate and morphology of bone fusion and rod fracture rate in the thoracic and lumbosacral vertebrae were evaluated. Patients were divided into the thoracic spine spontaneous bone fusion group and the bone fusion failure group. The data of various spinopelvic parameters and the incidence of complications were compared. The vertebral body conditions in the thoracic spine were classified as less degenerative (type N), osteophyte (type O), and diffuse idiopathic skeletal hyperostosis (DISH) (type D). Results: After three postoperative years, the bone fusion rates were 54%, 95%, and 89% for the thoracic, lumbar, and lumbosacral spine, respectively. Screw loosening in the thoracic vertebrae was significantly higher in the bone fusion failure group, while no significant differences were observed in the spinopelvic parameters, Oswestry Disability Index (ODI), and rate of proximal junctional kyphosis and rod fractures. Type N vertebral body condition and screw loosening were identified as risk factors for spontaneous bone fusion failure in the thoracic spine. Conclusion: This study indicated that spontaneous bone fusion is likely to be obtained without screw loosening, and even if bone fusion is not obtained, there is no effect on clinical results with the mid-term (3-year) results of CMIS without bone grafting in the thoracic spine.

Conditions for Achieving Postoperative Pelvic Incidence-Lumbar Lordosis < 10° in Circumferential Minimally Invasive Surgery for Adult Spinal Deformity

This retrospective study aimed to evaluate the clinical outcomes of circumferential minimally invasive surgery (CMIS) using lateral lumbar interbody fusion (LLIF) and percutaneous pedicle screw (PPS) in adult spinal deformity (ASD) patients, and to clarify the conditions for achieving postoperative pelvic incidence-lumbar lordosis (PI-LL) < 10°. Demographics and other parameters of ASD patients who underwent CMIS and who were divided into groups G (achieved postoperative PI-LL < 10°) and P (PI-LL ≥ 10°) were compared. Of the 145 included ASD patients who underwent CMIS, the average fused level, bleeding volume, operative time, and number of intervertebral discs that underwent LLIF were 10.3 ± 0.5 segments, 723 ± 375 mL, 366 ± 70 min, and 4.0 segments, respectively. The rod material was titanium alloy in all the cases. The PI-LL significantly improved from 37.3 ± 17.9° to 1.2 ± 12.2° postoperatively. Pre- and postoperative PI, postoperative LL, preoperative PI-LL, PI-LL after LLIF, and postoperative PI-LL were significantly larger in group P. PI-LL after LLIF was identified as a significant risk factor of postoperative PI-LL < 10° by logistic regression, and the cut-off value on receiver operating characteristic curve analysis was 20°. Sufficient correction was achieved by CMIS. If PI-LL after LLIF was ≤20°, it was corrected to the ideal alignment by the PPS procedure.

Threshold-based Monitoring of Compound Muscle Action Potentials for Percutaneous Pedicle Screw Placement in the Lumbosacral Spine: Can We Rely on Stimulation of the Uninsulated Screw to Provide a Valid Safety Warning?

STUDY DESIGN: A prospective analysis. OBJECTIVE: To test if threshold-based monitoring of compound muscle action potentials (CMAPs) by stimulating the screw loaded to uninsulated extender sleeve provides a valid safety warning for percutaneous pedicle screw (PPS) placements in the lumbosacral spine. SUMMARY OF BACKGROUND DATA: Utility of the CMAP monitoring to PPS procedures remains controversial. METHODS: A series of 202 patients underwent a total of 1664 lumbosacral PPS placements under CMAP monitoring without fluoroscopic guidance. The monitoring consisted of stimulating the PPS assembled to uninsulated extender sleeve and recording CMAPs from the vastus medialis, biceps femoris, tibialis anterior, and medial gastrocnemius. Automated steps of a threshold hunting algorithm using 0.2-ms duration pulses of increasing intensities delivered at 2/s allowed quick determination of a minimum stimulation current to evoke >100-µV amplitude CMAPs. RESULTS: At L2 through S1 spines, postoperative CT scans identified 51 medial or inferior pedicle wall breaches of 1536 screws (3.3%) without neurologic complications. The receiver operating characteristic curve analysis determined the critical cutoff threshold value of 27 mA (74% sensitivity and 95% specificity) for predicting 35 breaches of 627 screws (5.6%) at L2 and L3, and of 17 mA (100% sensitivity and 98% specificity) for 16 of 909 (1.8%) at L4 through S1. While advancing the screw, three breaches (5.9%) showed a particularly low threshold of ≤6-mA, allowing the surgeon to immediately redirect the screw and retest the new trajectory as safe. CONCLUSION: Screw stimulation with threshold hunting algorithm has a distinct advantage over the time-consuming insulated pilot hole stimulation, allowing an uninterrupted flow of the surgery. The present findings have documented practical usefulness and reliability of CMAP monitoring using direct stimulation of the PPS assembled to uninsulated extender sleeve.

A new technique useful for lumbosacral percutaneous pedicle screw placement without fluoroscopy or computer-aided navigation systems.

BACKGROUND: Percutaneous pedicle screw (PPS) placements in the lumbosacral spine generally rely on fluoroscopy at the expense of radiation exposure. Our accumulated experience in open PS placements without fluoroscopic guidance realized a consistent shift toward PPS insertion with newly developed devices, which require neither fluoroscopy nor navigation. We wish to report our new technique and evaluations of its accuracy. METHODS: Our equipment consisted of a pedicle targeting tool to identify and escort the cannulated awl to the correct starting point for cortical bone perforation and a cannulated awl-probe system with a guidewire to maintain the optimal position throughout the subsequent surgical steps. The surgeon could advance the blunt-tipped probe searching for the cancellous bone track using tactile feedback as experienced in open techniques. A 2-year period of transition from a free-hand (1169 screws in 286 patients) to the new PPS technique (1933 screws in 413 patients) allowed accuracy comparison between the two procedures using postoperative CT scans. RESULTS: Compared with the open-group, the PPS-group showed a lower rate of fully contained intrapedicular PS placements at L1 through S1, as a whole (90.7% vs 85.4%), but not at L4 through S1 (89.9% vs 90.2%). Less-accurate PPS placements at upper than lower lumbar spines in part reflect intended pedicle perforations laterally as a trade-off for avoiding facet violation immediately above the most cephalad screw. The PPS-group also had a higher incidence of PS-related transient nerve root complications (0% vs 1.7%). These values for the PPS-group, however, fell within those previously reported for free-hand or fluoroscopy techniques. CONCLUSIONS: Our new PPS technique, although useful for eliminating the potential risk of repeated radiation exposure, fell short of reaching the accuracy of the free-hand technique. Nerve integrity monitoring with PS stimulation, which we currently use, will help further improve the technical precision. STUDY DESIGN: Original Article. The study was approved by our institutional review boad (2,019,231).

Rod contour and overcorrection are risk factors of proximal junctional kyphosis after adult spinal deformity correction surgery.

PURPOSE: To examine the risk factors of proximal junctional kyphosis (PJK) after surgery for adult spinal deformity (ASD) focusing on rod contour. METHODS: Sixty-three patients with ASD who underwent surgery using lateral lumbar interbody fusion and percutaneous pedicle screws were analyzed. Fixation range was from the lower thoracic spine to the pelvis in all cases. Patients were divided into two groups. The PJK group consisted of 16 patients with PJK. The non-PJK group had 47 patients without PJK. We examined various spinopelvic parameters and parameters related to rod contour. RESULTS: Among the various spinal and pelvic parameters, those in the PJK group were significantly larger in terms of preoperative SVA and were significantly smaller in terms of postoperative "PI-LL." For parameters related to rod contour, the rod kyphotic curve at the thoracic spine in the PJK group was significantly less than that in the non-PJK group. The inclination of the pedicle screw at the upper instrumented vertebra (UIV) was significantly more cranial in the PJK group than in the non-PJK group. The kyphotic curve of the rod at the UIV was more parallel in the PJK group than in the non-PJK group. On logistic regression analysis, insufficient kyphotic curve at the thoracic spine along with UIV and overcorrection of the lumbar spine were identified as significant risk factors. CONCLUSIONS: Insufficient kyphotic curve of the rod in the thoracic spine along with UIV and overcorrection of the lumbar spine were noted as significant risk factors of PJK.

Usefulness of immunohistochemistry for the detection of the BRAF V600E mutation in Japanese lung adenocarcinoma.

PURPOSE: Mutations in components of the mitogen-activated protein kinase (MAPK) cascade may be a new candidate for target for lung cancer. The usefulness of immunohistochemistry (IHC) as a new approach for the detection of BRAF V600E in cancer patients has been recently reported. METHODS: To increase the sensitivity, we modified BRAF V600E expression detection assay by IHC using mutation specific antibody. From the screening step, we found a novel 599 insertion T BRAF mutation in lung adenocarcinoma. In this study included 26 surgically removed cases with EGFR, Kras, erbB2, EML4-ALK and KIF5B-RET wild-type (wt) lung adenocarcinomas, including 7 BRAF mutants (5 V600E, 1 N581I, and 1 novel 599 insertion T mutation) analyzed by DNA sequencing. Detection of the BRAF V600E mutation was carried out by the Dako EnVision™ FLEX detection system using the VE1 clone antibody and compared with the results of direct sequencing. RESULTS: The autostainer IHC VE1 assay was positive in 5 of 5 (100%) BRAF V600E-mutated tumors and negative in 20 of 21 (95.2%) BRAF non-V600E tumors, except for a novel 599 insertion T case. CONCLUSION: IHC using the VE1 clone and FLEX linker is a specific method for the detection BRAF V600E and may be an alternative to molecular biology for the detection of mutations in lung adenocarcinomas. This method might be useful for screening to use molecular target therapy for lung adenocarcinomas.

Nestin-positive hair follicle pluripotent stem cells can promote regeneration of impinged peripheral nerve injury.

Nestin-positive, keratin 15 (K15)-negative multipotent hair follicle stem cells are located above the hair follicle bulge. We have termed this location the hair follicle pluripotent stem cell area. We have previously shown that transplantation of nestin-expressing hair follicle stem cells can regenerate peripheral nerve and spinal cord injuries. In the present study, we regenerated the impinged sciatic nerve by transplanting hair follicle pluripotent stem cells. Human hair follicle stem cells were transplanted around the impinged sciatic nerve of ICR nude (nu/nu) mice. The hair follicle stem cells were transplanted between impinged sciatic nerve fragments of the mouse where they differentiated into glial fibrillary acidic protein-positive Schwann cells and promoted the recovery of pre-existing axons. The regenerated sciatic nerve functionally recovered. These multipotent hair follicle stem cells thereby provide a potential accessible, autologous source of stem cells for regeneration therapy of nerves degenerated by compression between bony or other hard surfaces.

RET expression and detection of KIF5B/RET gene rearrangements in Japanese lung cancer.

RET encodes the tyrosine kinase receptor of growth factors belonging to the glial-derived neurotrophic factor family. Recently, RET gene rearrangements with N-terminal of KIF5B gene were identified in lung adenocarcinomas from large-scale sequencing. We investigated RET mRNA expression by real-time reverse-transcriptase polymerase chain reaction (RT-PCR) assay using LightCycler, and KIF5B/RET gene rearrangements using newly established fluorescence in situ hybridization (FISH) analysis in surgically treated nonsmall cell lung cancer (NSCLC) cases. RET protein expression was also investigated by immunohistochemistry (IHC). This study included 157 surgically removed NSCLC cases for mRNA level analyses. The RET/ß actin mRNA levels were not significantly different between lung cancer (6.359 ± 15.268) and adjacent normal lung tissues (8.205 ± 28.931, P = 0.6332). Tumor/normal (T/N) ratio of RET/ß actin mRNA levels was not different within gender, stage, smoking status, and pathological subtypes. T/N ratio of RET/ß actin mRNA levels was significantly higher in KIF5B/RET rearrangement samples (161.763 ± 123.488) than in wild-type samples (5.9013 ± 17.148, P = 0.044). Although RET IHC positivity was not perfectly correlated with KIF5B/RET arrangement, we have detected the KIF5B/RET rearrangements using FISH analysis. Thus, we have successfully introduced FISH for diagnosing KIF5B/RET positive lung adenocarcinoma. This method facilitates the molecular evaluation for RET fusions and could be applicable in clinical practice to detect lung cancer that may be responsive to RET inhibitors.

NK cells are migrated and indispensable in the anti-tumor activity induced by CCL27 gene therapy.

Natural killer (NK) cells have been demonstrated could play an important role in the treatment of a number of tumors in mice. In the present study, chemokine CCL27, which be considered only selectively chemoattracts cutaneous lymphocyte antigen positive memory T cells and Langerhans cells, firstly demonstrated that it could induce the accumulation of NK cells into tumor by the intratumoral injection of CCL27-encoding fiber-mutant vector, AdRGD-CCL27. Experiments using spleen cell fractionation and RT-PCR showed CCL27 receptor, mCCR10, was strongly expressed in NK cells, suggesting the accumulation of NK cells in tumor was attributed to chemoattractant activity of CCL27 itself. Moreover, the combination of AdRGD-CCL27 and AdRGD-IL-12 induced the synergistic anti-tumor activity via NK-dependent manner and induced more NK cells infiltration into tumor nodule than that induced by AdRGD-CCL27 alone or AdRGD-IL-12 alone.

Combination of two fiber-mutant adenovirus vectors, one encoding the chemokine FKN and another encoding cytokine interleukin 12, elicits notably enhanced anti-tumor responses.

For achieving optimal cancer immunotherapy, it is anticipated that both the activation and infiltration of immune cells into tumor are indispensable. In the present study, fiber-mutant adenovirus vectors (Ad) encoding chemokine FKN, (AdRGD-FKN), and cytokine interleukin 12, (AdRGD-IL-12), were constructed. The in vivo gene expression of AdRGD was confirmed and the combination of both FKN and IL-12 encoding Ad elicited synergistic anti-tumor activity in ovarian carcinoma, which induced tumor regression in all tumor-bearing mice, while using FKN alone did not show notable tumor-suppressive effect. The treatment with both IL-12 and FKN induced long-term specific immunity against OV-HM tumors in tumor-rejected mice. The results of immunohistochemical staining for CD3+ and perforin-positive cells suggested that the failure of using FKN alone was because of the inactivation of infiltrated immune cells. In contrast, cotransduction with IL-12 and FKN could induce more activated tumor-infiltrating immune cells than that transducted with FKN or IL-12 alone. The results indicated that using both chemokine and cytokine might be a powerful tool and a promising way for effective cancer immunotherapy.

Tyrosine1248-phosphorylated HER2 expression and HER2 gene amplification in female invasive ductal carcinomas.

BACKGROUND: Phosphorylated HER2 (pHER2) may more accurately reflect the signaling and functional activity of the HER2 protein than detection of HER2 itself. The detection of HER2 gene amplification using fluorescence in situ hybridization (FISH) provides superior prognostic information for the diagnosis of breast cancer. However, the relationship between pHER2 expression in tissue samples and HER2 gene amplification remains unclear. METHODS: A total of 210 cases were recruited. The expression of HER2 and tyrosine (Tyr)1248-pHER2 was investigated by immunohistochemistry, and HER2 gene amplification was analyzed by FISH. Spearman's rank correlation test was employed to confirm correlation between HER2 and Tyr1248-pHER2. Chi-square and Student's t test were used to determine a significant difference between the baseline characteristics of tumors and the FISH, HER2 and Tyr1248-pHER2 results. The phosphorylation rate of HER2 was calculated using a digital-analysis system. RESULTS: HER2 expression was significantly (P < 0.001) associated with Tyr1248-pHER2 expression. HER2 gene amplification could be detected in 55 (26.2%) of the 210 tumors; 40 were HER2 positive and 32 were Tyr1248-pHER2 positive. The sensitivity and specificity of HER2 and Tyr1248-pHER2 for HER2 gene amplification were 72.7 and 58.2%, and 91.6 and 95.5%, respectively. In cases with an HER2 score of 2, and a phosphorylation score of 2 or 3, gene amplification was observed in 4 (80.0%) out of 5 tumors. CONCLUSIONS: Tyr1248-pHER2 expression is highly specific for HER2 gene amplification. The phosphorylation status might provide an adjunct to the assessment of gene amplification in patients with an HER2 score of 2.