mTORC1 directly phosphorylates and activates ERα upon estrogen stimulation.

Breast cancer is the leading cause of cancer-related deaths among women. Approximately 75% of breast cancers are estrogen receptor-α (ERα) positive, underscoring the dependence of cancer cells on estrogen for growth and survival. Patients treated with endocrine therapy often develop resistance, either de novo or acquired, which in some cases is caused by aberrations within the growth factor signaling pathways. The mechanistic target of rapamycin complex 1 (mTORC1) has emerged as a critical node in estrogenic signaling. We have previously shown that mTORC1 can phosphorylate and activate ERα on S167 via its effector-the 40S ribosomal S6 kinase 1 (S6K1). Presently, we have uncovered a direct link between mTORC1 and ERα. We found that ERα binds to regulatory-associated protein of mTOR (Raptor) and causes it to translocate to the nucleus upon estrogen stimulation. In addition, we identified mTOR as the kinase that phosphorylates ERα on S104/106 and activates transcription of ER target genes. Our findings show a direct link between mTORC1 and ERα, which further implicates mTORC1 signaling in the pathogenesis of ER-positive breast cancer and provides rationale for FDA-approved use of mTORC1 inhibitors in combination with endocrine agents for treatment of this disease.

Anterior cruciate ligament reconstruction. A prospective randomized study of three surgical methods.

A prospective randomized study was performed to determine the differences in results between three methods of anterior cruciate ligament reconstruction: autogenous bone-patellar tendon-bone graft (group 1), semitendinosus and gracilis tendon graft reconstruction combined with an extraarticular procedure (group 2), and semitendinosus and gracilis tendon graft reconstruction alone (group 3). Preoperatively, there were no significant differences between groups. At a mean of 35.4 +/- 11.6 months postoperatively, 102 patients returned for evaluation. International Knee Documentation Committee knee evaluation revealed no significant differences in symptoms, function, return to pre-injury activity, harvest site abnormalities, or limitation of motion between groups 1 and 3. Patients in group 2 had a higher incidence of patellofemoral crepitation and loss of motion than did patients in group 3. The mean manual maximum KT-1000 arthrometer side-to-side difference was 2.1 +/- 2.0 mm in group 1, which was statistically significantly better than the difference in group 3 (3.1 +/- 2.3 mm). Final knee rating showed that 34 of 35 patients in group 1, 23 of 34 patients in group 2, and 26 of 33 patients in group 3 had a normal or nearly normal overall knee rating. Anterior cruciate ligament reconstruction with a semitendinosus and gracilis or a patellar tendon autograft may yield similar subjective results; however, the patellar tendon autograft may provide better objective stability in the long term. In addition, there appears to be no benefit to combining an intraarticular anterior cruciate ligament reconstruction with an extraarticular procedure.

Anterior cruciate ligament reconstruction using the semitendinosus and gracilis tendons augmented by the losee iliotibial band tenodesis. A long-term study.

Seventy patients were evaluated after combined intra- and extraarticular reconstruction of the anterior cruciate ligament. Semitendinosus and gracilis tendons were used for the intraarticular reconstruction and Losee iliotibial band tenodesis for the extraarticular procedure. Minimum followup was 5 years (average, 7). Clinical examination and instrumented ligament examination with the KT-1000 arthrometer and the Cybex II dynamometer were used to evaluate results. Subjectively, patients had no complaints of instability with daily activities; 93% had no complaints of instability with athletic participation. Clinical examination demonstrated that 81% had less than a 1 + Lachman test; 98% had a negative pivot shift. Instrumented examination of the anteroposterior limits of motion revealed that only 57% had less than 3-mm side-to-side difference; 30% had 3- to 5-mm; and 13% had 6- to 9-mm. Mean hamstring muscle strength measured with the Cybex II dynamometer was 94% at 60 and 96% at 100 deg/sec. When evaluated with the Zarins scale, 90% were rated good or excellent, 6% fair, and 4% poor. This reconstructive procedure restores functional, but not necessarily normal, stability in most anterior cruciate ligament-deficient knees. It allows patients to increase activity levels without significant risk of additional injuries.

A novel method to differentiate between ping-pong and simultaneous exchange kinetics and its application to the anion exchanger of the HL60 cell.

We have developed a new test to differentiate between ping-pong and simultaneous mechanisms for tightly coupled anion exchange. This test requires the use of a dead-end reversible noncompetitive inhibitor. As an example, we have applied the test to the anion exchanger of the HL60 cell using the salicylic acid derivative 3,5-diiodosalicylic acid (DIS), which reversibly inhibits HL60 cell Cl/Cl exchange. The concentration of DIS that causes 50% inhibition (ID50) increased only slightly as either intra- or extracellular chloride was increased, indicating that DIS inhibits HL60 anion exchange in a noncompetitive manner. In agreement with this observation, plots of the slope of the Dixon plot as a function of 1/[Clo] or 1/[Cli] were fit with straight lines with nonzero intercepts, indicating that DIS does not compete with either of the substrates ([Clo] and [Cli]). The secondary Dixon slope test is based on the fact that, for a dead-end inhibitor such as DIS, the slope of the Dixon plot slope vs. 1/[Cli] (secondary Dixon slope or SDS) is independent of extracellular Cl when the exchange mechanism follows ping-pong kinetics. Similarly, the SDS calculated from a plot as a function of 1/[Clo] is also independent of intracellular Cl for a ping-pong exchanger. In contrast to this prediction, we found that for DIS inhibition of Cl/Cl exchange in HL60 cells the slope of the Dixon plot slope vs. 1/[Cli] decreased by a factor of 2.5-fold when [Clo] was increased from 1 to 11 mM (P < 0.0001). This change in the SDS rules out ping-pong kinetics, but is consistent with a simultaneous model of Cl/Cl exchange in which there are extra- and intracellular anion binding sites, both of which must be occupied by suitable anions in order to allow simultaneous exchange of the ions.

Instrumented evaluation of knee laxity: a comparison of five arthrometers.

We performed a prospective study on 50 subjects with normal knees and 50 patients with chronic unilateral disruption of the anterior cruciate ligament. In a randomized testing sequence, both groups were examined with five arthrometers: the MEDmetric KT-1000, the Stryker Knee Laxity Tester, the Acufex Knee Signature System, the Dyonics Dynamic Cruciate Tester, and the Genucom Knee Analysis System. Each examination was performed according to protocol with the knee at 30 degrees of flexion. The total anterior laxity measurements of the normal subjects using the Dyonics Dynamic Cruciate Tester and Acufex Knee Signature System were approximately half of the KT-1000, Stryker, and Genucom values. A comparison of the side-to-side measurements revealed no statistically significant difference in the values of the five arthrometers. However, the Genucom showed an unacceptably high number of normal subjects with laxity values that suggested an anterior cruciate ligament tear. Total anterior laxity measurements of the anterior cruciate ligament deficient knees were almost twice those of normal knees with each device except the Genucom. Comparison of the mean side-to-side difference in the patients with ACL deficiency demonstrated statistically significant differences in the values recorded with the five arthrometers. The side-to-side difference was greatest with the KT-1000 and least for the KSS. In the 89 N Lachman test, the KT-1000 and Stryker demonstrated the highest diagnostic accuracy. The maximum manual test improved the accuracy of each device. This study establishes the total anterior laxity measurements cannot be generalized from one device to another in either group of subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

Kinetics of DIDS inhibition of HL-60 cell anion exchange rules out ping-pong model with slippage.

According to the ping-pong model of band 3-mediated anion exchange, the transport protein has a single transport site, which can exist in either an inward-facing or an outward-facing conformation. Anions bind to these unloaded forms of the carrier, and translocation takes place only when a suitable anion is bound to the transport site. In a previous paper [Am. J. Physiol. 257 (Cell Physiol. 26): C520-C527, 1989], we had shown that the substrate kinetics of Cl-Cl exchange in the promyelocytic HL-60 cell cannot be explained by this simple ping-pong model of anion exchange but is consistent with a simultaneous model according to which both extracellular and intracellular anions must bind before simultaneous translocation can take place. In the present paper we show that external 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) inhibits anion exchange in HL-60 cells by competing with Cl- for binding to the outward-facing transport site. Furthermore, there is a linear dependence of the slope of the Dixon plot for inhibition by DIDS on the reciprocal of the intracellular Cl- concentration. This result clearly rules out a simple ping-pong scheme. In addition, the data also rule out a ping-pong model in which some translocation of the unloaded carrier is allowed (ping-pong model with slippage). The observed inhibition kinetics can be modeled by a simultaneous model of Cl-Cl exchange with competitive inhibition by DIDS.

Evaluation of hamstring strength following use of semitendinosus and gracilis tendons to reconstruct the anterior cruciate ligament.

Reconstruction of the anterior cruciate ligament using the semitendinosus and gracilis tendons combined with the appropriate extraarticular procedures has been performed by the authors in 482 cases. In 321 cases both the semitendinosus and gracilis tendons were used and in 161 the semitendinosus alone. This retrospective study was done to determine if the use of these two tendons resulted in any significant loss of hamstring strength. Evaluation of quadriceps strength was also done as a measure of postoperative rehabilitation. Fifty-one patients with an average follow-up time of 26.2 months were tested on the Cybex machine (Cybex Co., Ronkonkoma, New York) by two examiners using the same technique. The examiners had not participated in the surgery or rehabilitation of these patients. In the reconstructed knee in which both semitendinosus and gracilis were used, hamstring strength was found to average 99% compared to the normal knee. When the semitendinosus alone was used there was no difference (102%) from the normal knee. Quadriceps strength in both groups averaged 96% in comparison to the normal quadriceps. These results confirm that no significant loss of hamstring strength occurred when the semitendinosus and gracilis tendons were used to construct the anterior cruciate ligament. Further, quadriceps strength of 96% as compared to the normal knee indicated a very acceptable degree of postoperative rehabilitation in this series.

The relationship of tarsal coalitions to ankle sprains in athletes.

A review of charts and x-ray films of individuals who sustained ankle sprains was carried out to determine the presence of calcaneonavicular coalition. Coalitions were present in 63% of 215 patients with 223 ankle sprains. In 130 ankle sprains recorded as occurring directly in athletics, 65% of the x-ray films showed some degree of calcaneal navicular abnormality. This study found a much higher percentage of coalitions than anticipated in either the general population or the injured athletic population and reinforced the belief that there may be an underlying anatomic predisposition to ankle sprains.

The technique of cruciate ligament reconstruction.

Our technique of reconstruction of the anterior and posterior cruciate ligament in both acute and chronic tears, using both the semitendinosus and gracilis tendons, is described in detail. Combined rotatory instabilities were carefully evaluated and corrected during the same operation. Reefing of the posteromedial capsular ligament was done for anteromedial rotatory instability, and for significant valgus instability, the O'Donoghue medial reconstruction was performed. The Ellison procedure was initially done for associated anterolateral rotatory instability, but has been replaced by the Losee procedure. This combination of anterior cruciate ligament reconstruction with the appropriate extraarticular procedures has produced our most stable knee joints. Posterior cruciate ligament reconstruction was also performed through an anterior incision using both the semitendinosus and gracilis tendons. When both cruciates were found to be incompetent, the gracilis was used to reconstruct the posterior cruciate and the semitendinosus for the anterior cruciate. In addition, all associated rotatory instabilities were corrected at the same operation. This technique has been performed on 372 patients (342 anterior and 30 posterior cruciate ligament tears) by one surgeon over a period of five years. The average follow-up was 22 months. Objective comparison of the reconstructed knee with the sound knee showed 84% good results.