Does soft tissue balancing using intraoperative pressure sensors improve clinical outcomes in total knee arthroplasty? A protocol of a multicentre randomised controlled trial.

INTRODUCTION: Soft tissue imbalance is considered to be a major surgical cause of dissatisfaction following total knee arthroplasty (TKA). Surgeon-determined manual assessment of ligament tension has been shown to be a poor determinant of the true knee balance state. The recent introduction of intraoperative sensors, however, allows surgeons to precisely quantify knee compartment pressures and tibiofemoral kinematics, thereby optimising coronal and sagittal plane soft tissue balance. The primary hypothesis of this study is that achieving knee balance with use of sensors in TKA will improve patient-reported outcomes when compared with manual balancing. METHODS AND ANALYSIS: A multicentred, randomised controlled trial will compare patient-reported outcomes in 222 patients undergoing TKA using sensor-guided balancing versus manual balancing. The sensor will be used in both arms for purposes of data collection; however, surgeons will be blinded to the pressure data in patients randomised to manual balancing. The primary outcome will be the change from baseline to 1 year postoperatively in the mean of the four subscales of the Knee Injury and Osteoarthritis Outcome Score (KOOS4) that are most specific to TKA recovery: pain, symptoms, function and knee-related quality of life. Secondary outcomes will include the surgeon's capacity to determine knee balance, radiographic and functional measures and additional patient-reported outcomes. Normality of data will be assessed, and a Student's t-test and equivalent non-parametric tests will be used to compare differences in means among the two groups. ETHICS AND DISSEMINATION: Ethics approval was obtained from South Eastern Sydney Local Health District, Approval (HREC/18/POWH/320). Results of the trial will be presented at orthopaedic surgical meetings and submitted for publication in a peer-reviewed journal. TRIAL REGISTRATION NUMBER: ACTRN#12618000817246.

Does bone cement influence soft tissue balancing in total knee arthroplasty?

BACKGROUND: Soft tissue tension significantly affects the function of total knee arthroplasties. This study aims to evaluate if there is a difference in soft tissue tension, comparing trails to cemented definitive components in TKA. METHODS: We prospectively compared femorotibial compartment pressures before and after cement fixation of the components in 40 primary TKA. Femorotibial pressures were measured in the medial and lateral compartment with the knee in 10°, 45°, and 90° of flexion (six measurements per TKA), and the difference in pressure between both compartments was calculated in the three positions. RESULTS: The median femorotibial pressures were not significantly different following cement fixation. There was, however, a change in the difference between medial and lateral compartment pressures after cement fixation. The difference between both compartment pressures decreases after cement fixation. This difference is statistically significant only with the knee in 10° of flexion; mean (IQR) pressures change from 8.5 (five to 14) pounds to six (2.25-10) pounds (P = 0.01). CONCLUSION: Compartment pressures in TKA do not significantly change after cement fixation. The number of TKA that qualifies as 'balanced' increases after cement fixation, predominantly because the differences between the medial and lateral compartment pressures decrease.

Kinematic alignment technique for medial OXFORD UKA: An in-silico study.

BACKGROUND: Mobile bearing unicompartmental knee arthroplasty (UKA) Oxford™ components are recommended to be systematically and mechanically aligned (MA) for restoring the constitutional lower-limb alignment. Good long-term clinical outcomes have been generated with the medially implanted MA Oxford™, but some sub-optimal biomechanical-related complications still remain. Kinematic Alignment (KA) is a personalised technique for anatomically and kinematically implanting components (total knee, fixed bearing partial knee, total hip) aimed at creating more physiological prosthetic joint biomechanics. Interestingly, for decades the principles for implanting fixed bearing UKA components were consistent with those promoted by the KA technique, but differently formulated. We initiated this computational study to assess the feasibility of this technique with the Oxford™ components, as we thought this more anatomical implantation may be clinically advantageous. HYPOTHESIS: We surmised that kinematically aligning the Oxford™ medial UKA would maximise the prosthesis-bone interface through maximising the implants' size used (question 1), and alter, within an acceptable limit, the components' orientation (question 2) compared to conventional mechanical alignment. METHODS: A cohort of 40 consecutive medial osteoarthritic knee patients scheduled for UKA had a preoperative CT scan that was segmented to create 3D knee bone models. MA and KA of medial UKA Oxford® components (Zimmer-Biomet, Warsaw, Indiana, USA) were simulated. Component sizing and positioning were compared between the two techniques. RESULTS: We found no difference in component size, but significantly fewer occurrences of borderline fit with the KA simulation. KA technique oriented the femoral component 3.6° more valgus (from 1° varus to 7° valgus) and the tibial component 2.9° more varus (from 8° varus to 0°) compared to the MA technique. The tibial component slope in KA simulation was 6.4° posterior (from 0 to 12°) compared to a systematic 7° posterior for MA positioning. DISCUSSION AND CONCLUSION: Kinematic alignment of the medial Oxford™ generated a different, albeit still acceptable (Oxford group recommendations), implant orientation, in addition to a likely better shape-fit between components and the supportive bone cut, compared to the MA technique. The potential to improve the implants' interaction and to restore a more physiological bone loading makes the KA of Oxford™ an attractive, potentially clinically beneficial option. Clinical investigations are needed to assess its true value. LEVEL OF EVIDENCE: I, computational study.

Poor relationship between frontal tibiofemoral and trochlear anatomic parameters: Implications for designing a trochlea for kinematic alignment.

BACKGROUND: The kinematic alignment (KA) technique for total knee arthroplasty (TKA) is an emerging implant positioning philosophy that aims to restore constitutional knee anatomy to improve knee kinematics. At present, the KA technique aims to reconstruct native femorotibial (FT) joint alignment, however there is still insufficient consideration towards the inter-individual trochlear anatomy variability. Poor trochlear restoration may compromise clinical outcomes. Our study aimed at assessing the anatomical relationship between the native trochlea and other FT anatomical parameters. METHODS: Fifty-eight preoperative CT scans of low-grade knee arthritic patients were segmented to create 3D bone models. The FT and the PF anatomical parameters were measured using in-house software. Values were compared between different groups of lower limb and FT joint line (JL) orientation, and correlations between FT and PF anatomical parameters were assessed. RESULTS: We were unable to detect any significant correlation between groove orientation (frontal and axial) or groove radius and either the hip-knee-ankle (HKA), or the lateral distal femoral (LDFA), or the medial proximal tibial (MPTA), or the FTJL-mechanical axis (FTJLMAA) Angles. When considering the correlation within sub-groups of limb or JL orientation, we only found a positive correlation (r = 0.464, p = 0.022) in the varus lower limb (HKA ≤ 180°) sub-group between groove frontal orientation and LDFA. CONCLUSION: Our study shows that the determination of several limb, knee, and JL parameters is of poor value to predict individual trochlea anatomy. This raises the issue of how to improve femoral component design to achieve individualised FT and PF anatomical restoration with KATKA. LEVEL OF EVIDENCE: Level 1 - computational study.

The Reliability of Sensor-Assisted Soft Tissue Measurements in Primary Total Knee Arthroplasty.

BACKGROUND: In pursuit to improve soft tissue balancing in total knee arthroplasties (TKAs), a wireless device was introduced to assess femorotibial pressures. The aim of this study was to evaluate the reliability of this device. METHODS: After 33 TKAs were balanced by conventional techniques, contact pressures were measured using a wireless sensor 3 times in a row; twice while the examiner was blinded for the result (n = 29); and once while the examiner was able to see the result as visual feedback (n = 32). Femorotibial pressures were measured in the medial and lateral compartments with the knee in 10°, 45°, and 90° of flexion (6 measurements per TKA). Furthermore, both the combined pressure and the difference in pressure between the compartments was calculated throughout the 3 positions (together another 6 measurements per TKA). RESULTS: The intraclass correlation coefficient between the blind measurements was poor in 2 of the 12 (17%), moderate in 4 of 12 (33%), and good in 6 of 12 (50%) measurements. The intraclass correlation coefficient between the blind and observing measurement was poor in 2 of the 12 (17%), moderate in 6 of 12 (50%), and good in 4 of 12 (33%) measurements. Especially measurements in 10° of flexion are associated with poorer reliability. CONCLUSION: The wireless sensor has a moderate to good reliability in 83% of the measurements.

Precision insertion of percutaneous sacroiliac screws using a novel augmented reality-based navigation system: a pilot study.

PURPOSE: Augmented reality (AR) enables superimposition of virtual images onto the real world. The aim of this study is to present a novel AR-based navigation system for sacroiliac screw insertion and to evaluate its feasibility and accuracy in cadaveric experiments. METHODS: Six cadavers with intact pelvises were employed in our study. They were CT scanned and the pelvis and vessels were segmented into 3D models. The ideal trajectory of the sacroiliac screw was planned and represented visually as a cylinder. For the intervention, the head mounted display created a real-time AR environment by superimposing the virtual 3D models onto the surgeon's field of view. The screws were drilled into the pelvis as guided by the trajectory represented by the cylinder. Following the intervention, a repeat CT scan was performed to evaluate the accuracy of the system, by assessing the screw positions and the deviations between the planned trajectories and inserted screws. RESULTS: Post-operative CT images showed that all 12 screws were correctly placed with no perforation. The mean deviation between the planned trajectories and the inserted screws was 2.7 ± 1.2 mm at the bony entry point, 3.7 ± 1.1 mm at the screw tip, and the mean angular deviation between the two trajectories was 2.9° ± 1.1°. The mean deviation at the nerve root tunnels region on the sagittal plane was 3.6 ± 1.0 mm. CONCLUSIONS: This study suggests an intuitive approach for guiding screw placement by way of AR-based navigation. This approach was feasible and accurate. It may serve as a valuable tool for assisting percutaneous sacroiliac screw insertion in live surgery.

A morphometric study of normal and varus knees.

PURPOSE: The aim of the study was to investigate varus and normal knee morphologies to identify differences that may affect knee replacement alignment or design for varus knees. METHODS: Computed tomography scans of varus and normal knees were analyzed, and geometric shapes, points and axes were fit to the femur and tibia independently. These points were then projected in the three anatomical planes to measure the variations between the two groups. RESULTS: In the femur, varus knees had less femoral anteversion (p < 0.0001) and a larger medial extension facet (p < 0.05) compared with normal knees. In the tibia, the tubercle was found to be externally rotated in varus knees (12°), with a significant increase in the coronal slope (p = 0.001) and the extension facet angle (p = 0.002). CONCLUSIONS: The study highlighted the differences and similarities found between the two groups, which raises awareness on changes required during surgical intervention and component placement or design for a varus knee. This is particularly relevant for the design of patient-specific instrumentation and implants. LEVELS OF EVIDENCE: Diagnostic study, Level III.

Intra-operative reliability of ShapeMatch cutting guide placement in total knee arthroplasty.

Custom cutting guides based on pre-operative imaging have been introduced for total knee arthroplasty (TKA). The aim of this prospective cohort study was to assess the reliability of repeated placement of custom cutting guides by multiple surgeons in a group of patients undergoing TKA. Custom cutting guides (ShapeMatch®, Stryker Orthopaedics) were designed from pre-operative MRI scans. The treating surgeon placed each guide on the femur and tibia of each patient three times without pinning the block. The three-dimensional position and orientation of the guide was measured for each repetition using a computer navigation system. The surgeon was blinded to the navigation system display. Data from 24 patients and 6 surgeons were analyzed. Intraclass correlation coefficients for all measurement parameters were in the range 0.889-0.997 (excellent), and all comparisons were statistically significant (p < 0.001). The range for femoral varus/valgus was 0.0-1.5°, with 96% of patients being within 0.5°. For femoral flexion/extension the range was 0.0-3.5° (92% within 2.5°). On the tibia, varus/valgus had a range of 0.0-1.0° (92% within 0.5°), and for slope the range was 0.0-3.5° (92% within 2.5°). The high degree of agreement indicated that intra-surgeon variation was minimal and that the technique is reliable.

Bone morphogenetic protein-7 accelerates fracture healing in osteoporotic rats.

BACKGROUND: Osteoporosis is characterized by low bone mass, bone fragility and increased susceptibility to fracture. Fracture healing in osteoporosis is delayed and rates of implant failure are high with few biological treatment options available. This study aimed to determine whether a single dose of bone morphogenetic protein-7 (BMP-7) in a collagen/carboxy-methyl cellulose (CMC) composite enhanced fracture healing in an osteoporotic rat model. MATERIALS AND METHODS: An open femoral midshaft osteotomy was performed in female rats 3 months post-ovarectomy. Rats were randomized to receive either BMP-7 composite (n = 30) or composite alone (n = 30) at the fracture site during surgery. Thereafter calluses were collected on days 12, 20 and 31. Callus cross-sectional area, bone mineral density, biomechanical stiffness and maximum torque, radiographic bony union and histological callus maturity were evaluated at each time point. RESULTS: There were statistically significant increases in bone mineral density and callus cross-section area at all time points in the BMP-7 group as compared to controls and biomechanical readings showed stronger bones at day 31 in the BMP-7 group. Histological and radiographic evaluation indicated significant acceleration of bony union in the BMP-7 group as compared to controls. CONCLUSION: This study demonstrated that BMP-7 accelerates fracture healing in an oestrogen-deficient environment in a rat femoral fracture healing model to scientific relevance level I. The use of BMP-7 composite could offer orthopedic surgeons an advantage over oestrogen therapy, enhancing osteoporotic fracture healing with a single, locally applied dose at the time of surgery, potentially overcoming delays in healing caused by the osteoporotic state.

The changing role of pathology in breast cancer diagnosis and treatment.

Pathological examination has been the gold standard for diagnosis in cancer and its role has also included the elucidation of etiology, pathogenesis, clinicopathological correlation, and prognostication. The advent of newer technologies and the realization that breast cancer is heterogeneous has shifted the focus to prognostication, with increased attention being paid to the identification of morphological features and immunohistochemical markers of prognostic relevance. However, despite the massive efforts invested in the identification of immunohistochemical biomarkers in breast cancer the majority have not proven to be of value in multivariate analyses and only estrogen receptor, progesterone receptor, and Her2/neu expression have remained essential components of pathological examination. These 3 markers were initially employed for prognostication but their role in treatment also rendered them of predictive value. Newer molecular methods, especially high-throughput technologies, have shown that even morphologically similar subtypes of breast cancer can show molecular heterogeneity; moreover, infiltrating ductal carcinoma can be separated into at least 4 molecular subtypes designated luminal (ER+, PR+, and Her2/neu-), Her2 overexpressing (ER-, PR-, and Her2/neu+), basal-like (ER-, PR-, Her2/neu-, and CK5/6+, EGFR+), and normal breast-like (ER-, PR-, and Her2/neu-), each with different clinical outcomes. The importance of proliferative gene expression in these subtypes has been demonstrated and surrogate immunohistochemical markers include ER, PR, Her2/neu, and Ki67 for the more expensive molecular tests. Molecular technologies, importantly, have not only provided further insights into the heterogeneity of breast cancer but have also opened new avenues for treatment through the identification of signaling molecules important in the proliferation and survival of the neoplastic cells. The treatment of cancer thus shifts from the conventional approach of 'one size fits all' to one of personalized treatment tailored to the specific characteristics of the tumor. Pathologists continue to play their traditional role in diagnosis but, as purveyors of the excised tissue, pathologists now have the additional role of identifying biomarkers responsive to therapeutic manipulation, thus playing an inextricable role as diagnostic oncologists in the management of breast cancer.

Standardization in immunohistology.

The rapid acceptance of immunohistology as an invaluable adjunct to morphologic diagnosis has been possible because of the development of new and more sensitive antibodies and detection systems that allow its application to formalin-fixed, paraffin-embedded tissue (FFPT). More importantly, antigen-retrieval techniques have resulted in some degree of consistency allowing immunohistology to be used reliably as a diagnostic tool. The advent of prognostic and predictive biomarkers, and the desire for individualized therapy has resulted in mounting pressure to employ the immunohistological assay in a quantitative manner. While it was not a major issue when the technique was employed in a qualitative manner, the numerous variables in the preanalytical and analytical phases of the test procedure that influence the immunoexpression of proteins in FFPT become critical to standardization. Tissue fixation is pivotal to antigen preservation but exposure to fixative prior to accessioning by the laboratory is not controlled. Antigen retrieval, crucial in the analytical phase, continues to be employed in an empirical manner with the actual mechanism of action remaining elusive. There is great variation in reagents, methodology, and duration of tissue processing and immunostaining procedure, and the detection systems employed are not standardized between laboratories. While many of these variables are offset by the application of antigen retrieval, which enables the detection of a wide range of antigens in FFPT, the method itself is not standardized. This myriad of variables makes it inappropriate to provide meaningful comparisons of results obtained in different laboratories and even in the same laboratory, as in current practice, each specimen experiences different preanalytical variables. Furthermore, variables in interpretation exist and cutoff thresholds for positivity differ. Failure to recognize false-positive and false-negative stains leads to further errors of quantitative measurement. Many of the problems relating to the technology and interpretation of immunostaining originate from failure to recognize that this procedure is different from other histological stains and involves many more steps that cannot be monitored until the end result is attained. While several remedial measures can be suggested to address some of these problems, accurate and reproducible quantitative assessment of immunostains presently remains elusive as important variables that impact on antigen preservation in the paraffin-embedded biopsy -cannot be standardized.

Microwaves for chromogenic in situ hybridization.

In situ hybridization can be employed in formalin-fixed, paraffin-embedded tissue sections (FFPT) and allows direct visualization of amplified genes and chromosomes in individual cell nuclei. Fluorescence in situ hybridization (FISH) is the most widely employed method, but the fluorescence preparations suffer from the main disadvantages of fading over time and poor visualization, the latter making it difficult to accurately separate invasive from in situ cancer cells. Chromogenic in situ hybridization (CISH) is a viable alternative to FISH in FFPT as it employs a peroxidase reaction to visualize the chromogen thus allowing the convenience of bright field microscopy and the correlation of the visualized gene amplification with cytomorphology. It is relatively less expensive and allows a permanent record, with several studies attesting to its validity. As with FISH, heat pretreatment and enzyme digestion are two critical components of the protocol. We describe a protocol for CISH in which a microwave-induced target retrieval step is introduced as a replacement for heat pretreatment. The same procedure is performed following enzyme digestion to produce consistent signals in amplified and nonamplified cells that are both larger in size and numbers when compared with those produced by the conventional protocol.

Immunohistology--past, present, and future.

The rapid development of immunohistochemistry, a morphology-based technique, has come about through refinements in detection systems and an increasing range of sensitive and specific antibodies that have allowed application of the technique to formalin-fixed, paraffin-embedded tissues. The introduction of heat-induced antigen retrieval has been a significant milestone to compliment these developments so that the immunohistochemistry is firmly entrenched as an indispensable adjunct to morphologic diagnosis. Although this ancillary stain was initially used in a qualitative manner, problems surrounding the many variables that influence antigen preservation in formalin-fixed, paraffin-embedded tissues were not a major issue and laboratories strived to optimize their staining protocols to the material they accessioned and processed. The advent of personalized medicine and targeted cancer treatment has imposed the need to quantitate the stain reaction product and has resulted in calls to standardize the process of immunostaining. A closer examination of the variables that influence the ability to show antigens in formalin-fixed, paraffin-embedded tissues revealed many important variables, particularly in the preanalytical phase of the assay, that are beyond the control of the accessioning laboratory. Although analytical factors have the potential to be standardized, the actions of many pivotal procedures including fixation and antigen retrieval are not completely understood. Postanalytical processes including threshold and cut-off values require consensus and standardization and it is clear that some of these goals can be achieved through the direction of national and international organizations associated with cancer diagnosis and treatment. With the ability to serve as a surrogate marker of many genetic abnormalities, immunohistochemistry enters a new era and the need to better understand some of the mechanisms fundamental to the technique become more pressing and the development of true quantitative assays is imperative. There is also an increasing appreciation that the technique highlights patterns of staining that reflect exquisite localization to organelles and tissue structures that are not appreciable in routine stains, adding a further dimension to morphologic diagnosis.

BMP-7 in combination with estrogen enhances bone formation in a fracture callus explant culture.

In postmenopausal women, estrogen withdrawal results in decrease in bone density or osteoporosis. Osteoporosis leads to fracture and retards bone-healing response. Bone morphogenetic protein-7 (BMP-7), a member of the transforming-growth factor-beta superfamily, has been shown as a promising candidate that stimulates bone growth in its application to fracture healing. The purpose of this study was to determine whether BMP-7 could enhance bone formation in the absence of estrogen. Female rats underwent a controlled closed fracture at the midshaft of the right femur. The callus tissues were harvested from the fracture site eight days following the fracture, and were cultured in serum-free media. The explanted callus tissues were then treated with BMP-7, estrogen (E2) or both. We assessed bone formation by measuring alkaline phosphatase (AP) activity, expression of an osteogenic transcription factor, Runt-related transcription factor-2 (Runx2), production of nitric oxide (NO), and calcium mineralization. Supplementation of serum-free cultures with BMP-7 alone increased cell proliferation by twofold, caused a 6.5-fold increase in AP activity, and enhanced calcium mineralization after 48 h. Moreover, BMP-7 in combination with E2 caused a 8.2-fold increase in the AP activity. Runx2 protein expression was increased following stimulation with BMP-7 and E2. Interestingly, E2 induced the amount of NO production by twofold, whereas BMP-7 did not, either alone or with E2. Thus, BMP-7 could enhance early and late markers of bone fracture healing in callus explant cultures, except for NO. BMP-7 could be a promising growth factor in the treatment of fractures as a consequence of osteoporosis.

Citraconic anhydride: a new antigen retrieval solution.

BACKGROUND: The introduction of heat-induced antigen retrieval has been a major milestone in diagnostic immunohistochemistry, enabling the application of many antibodies to fixed paraffin-embedded tissue sections. A number of important variables affect the preservation of tissue antigens, among which are analytical variables including the antigen retrieval methodology. Temperature of retrieval, duration of heating, source of heat, pH and nature of retrieval solution are among more important variables pivotal to results. Citrate buffer at pH 6.0 has become widely embraced as the universal fixative but some antibodies remain capricious and yield poor staining results. METHODS: This study examines the recent suggestion that citraconic anhydride may be a suitable universal retrieval reagent. Immunostaining of 65 commonly employed antibodies following microwave antigen retrieval in 0.05% citraconic anhydride for 10 minutes at 98 degrees C was compared with consecutive tissues sections subjected to antigen retrieval in citrate buffer at pH 6.0 at the same duration and temperature. RESULTS: Thirty-five of the 65 antibodies examined yielded more intense staining following antigen retrieval in citraconic anhydride, including some capricious antibodies such as MyoD1, myogenin, perforin, TIA-1, Tdt, RET and MiTF, confirming the efficacy of this retrieval solution. CONCLUSION: It is recommended that consideration be given to 0.05% citraconic anhydride as an antigen retrieval solution, particularly for antibodies that fail to work or stain weakly in fixed paraffin-embedded tissue sections.

Glomeruloid peritoneal implants in ovarian serous borderline tumours--distinction between invasive and non-invasive implants and pathogenesis.

AIMS: To determine whether or not the glomeruloid implants (GI) composed of papillary cores within clear spaces lined by mesothelial cells or tumour cells located in superficial or deep peritoneal tissue in ovarian serous borderline tumours (SBTs) are invasive. METHODS AND RESULTS: We examined the differences in incidence, histological and immunohistochemical findings among three groups: 100 GI with mesothelial cells lining clear space (type I), 100 GI with tumour cells lining clear space (type II), and 100 invasive implants with clefts but no lining cells from 30 cases of SBT with peritoneal implants. The type I lesion had characteristics of non-invasive implants with a tendency for smooth contours (100/100), superficial location (71/100), absence of desmoplasia (100/100) and absence of surrounding destructive invasion (100/100), In contrast, type II GI had irregular contours (67/100), deep location (93/100), presence of desmoplastic reaction (100/100) and presence of destructive invasion (12/100). Immunohistological studies suggested intermediate forms between the two types of lesions. CONCLUSIONS: Type I GI are non-invasive implants, whereas type II GI are invasive implants and it is important to evaluate the presence and nature of cells lining the clear space in determining whether implants associated with ovarian SBTs are invasive or not.

p16, cyclin D1, Ki-67, and AMACR as markers for dysplasia in Barrett esophagus.

Barrett esophagus (BE) is an established precursor of esophageal adenocarcinoma (AdenoCa). One hundred and one cases of BE diagnosed by esophageal biopsy and resections were examined morphologically for dysplasia. These were categorized as BE without dysplasia (n=25), indefinite for dysplasia (IND, n=17), low-grade dysplasia (LGD, n=18), high-grade dysplasia (HGD, n=15), and AdenoCa (n=26). Immunostaining for p16 (INK4A/CDKN2A), Cyclin D1 (CCND1), Ki-67, and alpha-methylacyl-CoA racemase (AMACR) was employed to assess their potential as diagnostic discriminators. Abnormal p16 expression (negative, cytoplasmic, or combined cytoplasmic and nuclear staining) was present in all categories, rising from 68% in BE without dysplasia to 100% in AdenoCa, with cytoplasmic staining only showing a significant correlation with the severity of dysplasia. Cyclin D1 expression was present in almost all cases, but high expression (>50% cells positive) was displayed mostly in HGD and AdenoCa (46.7% and 42.3%, respectively). Ki-67 index increased with the severity of dysplasia and labeling extended from the lower third of the crypts to the superficial epithelium. The frequency of AMACR-positivity was 12% in BE, 47.1% in IND, 44.4% in LGD, 93.3% in HGD, and 96.2% in AdenoCa. The intensity and extent of AMACR staining also increased with the severity of dysplasia. Aberrant p16 and high-cyclin D1 expression may reflect early genetic events during the progression of Barrett-associated carcinogenesis. Cytoplasmic staining of p16 is specific. It may represent a different pathway of p16 dysfunction. The pattern and extent of Ki-67 staining and AMACR overexpression are useful additional parameters for identifying dysplasia in BE.

CDX2 and villin are useful markers of intestinal metaplasia in the diagnosis of Barrett esophagus.

The identification of intestinal metaplasia (IM) in the esophagus is necessary for the selection of patients with Barrett esophagus (BE) for surveillance. We studied 108 esophageal biopsy and resection specimens, clinically diagnosed as BE, and stained them for CDX2, villin, HepPar-1, and cytokeratin (CK) 7 to investigate sensitivity for identifying IM. H&E-stained sections showed definite goblet cells in 94 cases. CDX2 and villin were positive in all 94 cases. Of 38 cardia- and 9 fundic-type mucosa samples associated with BE, 13 (34%) and 0 (0%) displayed CDX2 positivity and 21 (55%) and 1 (11%) displayed villin positivity, respectively. HepPar-1 was positive in 54 (57%) of 94 cases with IM and negative in the associated cardia- and fundic-type mucosa. A full-thickness CK7 staining pattern was present in 90 (96%) of samples with IM and 22 (58%) and 0 (0%) of the associated cardia- and fundic-type mucosa, respectively. None of 20 control samples of morphologically normal gastric mucosa stained for CDX2 or villin. CDX2 and villin are sensitive markers for early-stage IM and can supplement the histologic identification of this premalignant condition in the esophagus.