Unbiased global optimization of Lennard-Jones clusters for N < or =201 using the conformational space annealing method.
We apply the conformational space annealing method to the Lennard-Jones clusters and find all known lowest energy configurations up to 201 atoms, without using extra information of the problem such as the structures of the known global energy minima. In addition, the robustness of the algorithm with respect to the randomness of initial conditions of the problem is demonstrated by ten successful independent runs up to 183 atoms. Our results indicate that this method is a general and yet efficient global optimization algorithm applicable to many systems.

title: Protein structure prediction based on fragment assembly and parameter optimization.
abstract: We propose a novel method for ab-initio prediction of protein tertiary structures based on the fragment assembly and global optimization. Fifteen residue long fragment libraries are constructed using the secondary structure prediction method PREDICT, and fragments in these libraries are assembled to generate full-length chains of a query protein. Tertiary structures of 50 to 100 conformations are obtained by minimizing an energy function for proteins, using the conformational space annealing method that enables one to sample diverse low-lying local minima of the energy. Then in order to enhance the performance of the prediction method, we optimize the linear parameters of the energy function, so that the native-like conformations become energetically more favorable than the non-native ones for proteins with known structures. We test the feasibility of the parameter optimization procedure by applying it to the training set consisting of three proteins: the 10-55 residue fragment of staphylococcal protein A (PDB ID 1bdd), a designed protein betanova, and 1fsd.

title: Measures for the assessment of fuzzy predictions of protein secondary structure.
abstract: Many of the recent secondary structure prediction methods incorporate the idea of fuzzy set theory, where instead of assigning a definite secondary structure to a query residue, probability for the residue being in each of the conformational states is estimated. Moreover, continuous assignment of conformational states to the experimentally observed protein structures can be performed in order to reflect inherent flexibility. Although various measures have been developed for evaluating performances of secondary structure prediction methods, they depend only on the most probable secondary structures. They do not assess the accuracy of the probabilities produced by fuzzy prediction methods, and they cannot incorporate information contained in continuous assignments of conformational states to observed structures. Three important measures for evaluating performance of a secondary structure prediction algorithm, Q score, Segment OVerlap (SOV) measure, and the k-state correlation coefficient (Corr), are deformed into fuzzy measures F score, Fuzzy OVerlap (FOV) measure, and the fuzzy correlation coefficient (Forr), so that the new measures not only assess probabilistic outputs of fuzzy prediction methods, but also incorporate information from continuous assignments of secondary structure. As an example of application, prediction results of four fuzzy secondary structure prediction methods, PSIPRED, PROFking, SABLE, and PREDICT, are assessed using the new fuzzy measures.

title: Microcanonical origin of the maximum entropy principle for open systems.
abstract: There are two distinct approaches for deriving the canonical ensemble. The canonical ensemble either follows as a special limit of the microcanonical ensemble or alternatively follows from the maximum entropy principle. We show the equivalence of these two approaches by applying the maximum entropy formulation to a closed universe consisting of an open system plus bath. We show that the target function for deriving the canonical distribution emerges as a natural consequence of partial maximization of the entropy over the bath degrees of freedom alone. By extending this mathematical formalism to dynamical paths rather than equilibrium ensembles, the result provides an alternative justification for the principle of path entropy maximization as well.

title: Protein loop modeling by using fragment assembly and analytical loop closure.
abstract: Protein loops are often involved in important biological functions such as molecular recognition, signal transduction, or enzymatic action. The three dimensional structures of loops can provide essential information for understanding molecular mechanisms behind protein functions. In this article, we develop a novel method for protein loop modeling, where the loop conformations are generated by fragment assembly and analytical loop closure. The fragment assembly method reduces the conformational space drastically, and the analytical loop closure method finds the geometrically consistent loop conformations efficiently. We also derive an analytic formula for the gradient of any analytical function of dihedral angles in the space of closed loops. The gradient can be used to optimize various restraints derived from experiments or databases, for example restraints for preferential interactions between specific residues or for preferred backbone angles. We demonstrate that the current loop modeling method outperforms previous methods that employ residue-based torsion angle maps or different loop closure strategies when tested on two sets of loop targets of lengths ranging from 4 to 12.

title: Prediction of protein tertiary structure using PROFESY, a novel method based on fragment assembly and conformational space annealing.
abstract: A novel method for ab initio prediction of protein tertiary structures, PROFESY (PROFile Enumerating SYstem), is proposed. This method utilizes the secondary structure prediction information of a query sequence and the fragment assembly procedure based on global optimization. Fifteen-residue-long fragment libraries are constructed using the secondary structure prediction method PREDICT, and fragments in these libraries are assembled to generate full-length chains of a query protein. Tertiary structures of 50 to 100 conformations are obtained by minimizing an energy function for proteins, using the conformational space annealing method that enables one to sample diverse low-lying local minima of the energy. We apply PROFESY for benchmark tests to proteins with known structures to demonstrate its feasibility. In addition, we participated in CASP5 and applied PROFESY to four new-fold targets for blind prediction. The results are quite promising, despite the fact that PROFESY was in its early stages of development. In particular, PROFESY successfully provided us the best model-one structure for the target T0161.

title: Conditional deletion of the focal adhesion kinase FAK alters remodeling of the blood-brain barrier in glioma.
abstract: Gliomas generally infiltrate the surrounding normal brain parenchyma, a process associated with increased vascular permeability (VP) and dysregulation of the blood-brain barrier (BBB). However, the molecular mechanisms underlying glioma-induced VP in the brain remain poorly understood. Using a conditional, endothelium-specific deletion of the focal adhesion kinase (FAK) in the mouse (FAK CKO), we show that FAK is critical for destabilization of the tumor endothelium in tumor-bearing mice, with mutant mice exhibiting a relatively normalized vasculature compared with wild-type mice (FAK WT). Tumor vessels in the FAK CKO mice displayed reduced VP compared with FAK WT mice, resulting in reduced tumor growth. Additionally, FAK CKO mice displayed partial restoration of cell-cell junction proteins in the tumor vessels and astrocyte-endothelium interactions in tumors, revealing an additional role of astrocytes in mediating tumor-induced VP. Together, these results provide genetic evidence that FAK is a mediator of tumor-induced VP in the brain. Our findings may help understand how therapeutics might be used to regulate specific cell-type interactions to restore BBB structure/function in cancer and perhaps other pathologic conditions.

title: Motion correction for phase-resolved dynamic optical coherence tomography imaging of rodent cerebral cortex
abstract: Cardiac and respiratory motions in animals are the primary source of image quality degradation in dynamic imaging studies, especially when using phase-resolved imaging modalities such as spectral-domain optical coherence tomography (SD-OCT), whose phase signal is very sensitive to movements of the sample. This study demonstrates a method with which to compensate for motion artifacts in dynamic SD-OCT imaging of the rodent cerebral cortex. We observed that respiratory and cardiac motions mainly caused, respectively, bulk image shifts (BISs) and global phase fluctuations (GPFs). A cross-correlation maximization-based shift correction algorithm was effective in suppressing BISs, while GPFs were significantly reduced by removing axial and lateral global phase variations. In addition, a non-origin-centered GPF correction algorithm was examined. Several combinations of these algorithms were tested to find an optimized approach that improved image stability from 0.5 to 0.8 in terms of the cross-correlation over 4 s of dynamic imaging, and reduced phase noise by two orders of magnitude in ~8% voxels.

title: Underlying venous pathology causing perimesencephalic subarachnoid hemorrhage.
abstract: Perimesencephalic subarachnoid hemorrhage (PSH) is a relatively benign clinical entity with a low risk of recurrent bleeding. The precise etiology of PSH has not yet been determined. We report here three cases of PSH with clinical and radiological features that support a venous system as a cause.



The first patient, a 72-year-old woman, had PSH and venous hemorrhagic infarct in the left thalamus on non-contrast CT. Subsequent cerebral angiography revealed widespread thrombosis in the cerebral venous system, a potential cause for reflux overflow hemorrhage. The second patient, a 55-year-old man with an established diagnosis of neuro-Behçet's disease, a well-known cause for cerebral venulitis, presented with PSH one year later. The third patient, a 39-year-old female, with incomplete Behçet's disease was admitted with PSH.



Current concepts on the anatomic origin and the possible pathophysiologic mechanism leading to PSH are discussed. The underlying pathological conditions in the venous system in our cases provide theoretical clues to the anatomic origin of PSH in general.

title: Rapid determination of bacteria in pools.
abstract: If a method for rapid determination of bacterial levels in swimming pools and whirlpools were available, bacterial health risks could be avoided before exposure occurred. Such a method would enable public-health authorities to make timely decisions about closing or reopening pools and to adjust disinfectant doses. The purpose of this study was to determine if a rapid adenosine triphosphate (ATP) bioluminescence method would serve this purpose. Bacteriological analyses were completed on samples from swimming pools and whirlpools in Washtenaw County, Michigan. The results from the ATP assay were compared with those from the conventional plate count method and were found to be highly correlated (r = .92). To assess whether a prefiltration step was necessary to delete nonbacterial ATP--which might originate from bathers' skin cells or other debris--two procedures were tested: one with prefiltration and one without prefiltration. The results for prefiltered samples were not statistically different from the results for non-prefiltered samples (p > .05), for both the heterotrophic plate counts and the ATP bioluminescence method. This study indicates that the ATP assay can estimate bacterial levels in pools within minutes and without prefiltration.

title: Murine vibrissae cultured in serum-free medium reinitiate anagen.
abstract: Aberrant regulation of the hair cycle has been implicated in human boldness (Reviewed in (Nakamura et al, 2001; Paus and Cotsarelis, 1999)). Although several modulators of mammalian hair follicle cycle have been recently described, discovery of effective treatment for boldness has suffered from the absence of a reliable in vitro culture systems in which the hair cycle can be assessed easily and inexpensively (Stenn and Paus, 2001). Several laboratories have established serum-free culture system where vibrissa can grow and differentiate in vitro (Supplemental Table 1, (Jindo et al, 1993; Robinson et al, 1997; Yano et al, 2001). Although rat vibrissae cultured for 23 days were reported to share histological similarities with catagen or pro-anagen (telogen) stage follicles (Philpott and Kealey, 2000), these follicles did not show any progress beyond pro-anagen phase nor did they produce a new hair shaft (Philpott and Kealey, 2000). In this report, we demonstrate that a simple modification permits murine vibrissae in the current in vitro culture system to reinitiate anagen. This will accelerate the development of screening systems aimed at modifying the hair cycle. 
 
To establish a modified in vitro vibrissa culture system, anagen-stage vibrissae were carefully isolated from 14-day-old mice and the tip of the vibrissa shafts were anchored in a stripe of sterilized silicone grease placed on the culture dish through a 3 ml syringe. The plate was then filled with serum-free medium (Figure 1a, Supplemental Material and Method). Out of 86 cultured vibrissae collected from three pups, 81 vibrissae (94%) showed measurable shaft growth (Table 1, Figure 1b). Two of 81 vibrissae were lost, and 17 developed abnormal (kinked) fiber and were omitted from growth measurement. Of the remaining 62 vibrissae, straight shafts were produced by all at a rate of 0.3–0.5mm/day for the first three days. While some follicles maintained this growth rate through the fifth day of culture (Figure 1b and 1c and data not shown), others began a gradual decline in growth rate (Figure 1b and 1c, ​,2a2a and data not shown). As previously reported (Jindo et al, 1993; Robinson et al, 1997), growth rates of all follicles slowed down considerably after 5 days in culture, indicating independence from culture conditions (Supplemental Table 1). 
 
 
 
Figure 1 
 
Vibrissae produce hair shafts and reinitiate anagen in serum-free culture medium 
 
 
 
 
 
Figure 2 
 
Histological and immunohistochemical characteristics of new anagen follicles 
 
 
 
 
 
Table 1 
 
Summary of the in vitro vibrissa culture 
 
 
 
Hair shaft elongation rate decreases and eventually stops when follicles enter catagen/telogen phase in vivo (Alonso and Fuchs, 2006). Several days after cessation of hair shaft growth, cultured vibrissae follicles continuously changed their morphology, at times visible through their collagen capsules . Strikingly, dissection and removal of collagen capsules from 21-day-cultured vibrissae revealed that all vibrissae produced a second shaft (Figures 1d and 2a–h, arrows; Figure S1), readily distinguishable from the original shaft whose proximal end formed club-end indicative of a completed catagen (Figures 1 and ​and22 closed arrowheads). Histological and immunohistochemical analyses using AE13, a marker for cortex/cuticle specific keratins, confirmed the formation of new hair shafts (Figures 2c–f and S1). The new shafts were produced in proximal bulbs that appeared smaller in size than the original ones before culture. Since the original hair growth stopped within the first 5 days of culture, and since a new shaft emerges from a secondary bulb, we concluded that vibrissa follicles from CD1 mice reinitiated anagen phase by regenerating hair bulbs . Staining for alkaline phosphatase activity identified an intact (but small) dermal papilla within the new bulb ; Young and Oliver, 1976). Moreover, the hair matrix cells were positive for Ki-67, a proliferation marker, even after 13 days in culture (Figure 2h), indicating that the formation of new shaft was the result of active proliferation and differentiation of matrix cells in cultured vibrissae. Similar cycling patterns were also observed with vibrissae from 14-day-old C57BL/6 mice , indicating that hair cycle in vitro is independent of mouse strains. 
 
To better visualize the hair cycle process, we next performed vibrissae culture after careful removal of their collagen capsules . Within 2 days, the proximal end of the vibrissa keratinized, forming a club hair, at which time dermal papilla structures were no longer discernable (Figure 2i and data not shown). A day later, however, a bud-like structure emerged from the proximal part of the bulb in eight of ten cultured vibrissae. Of these, one vibrissa continued growing outwards, eventually forming a new bulb producing a vibrissa thinner than the club hair . Strikingly, we observed that this vibrissa appeared to enter a third anagen, producing a new bud from the second bulb . Collectively, the data presented here indicate that in our in vitro culture system, vibrissa follicles enter a second anagen phase and it may be possible for a few to enter a third cycle in culture. 
 
We are not certain which modification in our culture system permits vibrissae to cycle. We do not think this is caused by differences in media composition, mouse strain, or frequency of media change. Our attempt to grow vibrissae at the air-fluid interface as in (Jindo et al, 1993) showed that one of five vibrissae formed a new hair shaft (Figure S3, Arrows), indicating that hair cycle is not strictly dependent on culture media or on the frequency of media change. However, we observed that on a solid substrate most vibrissae gradually lost their hair matrix possibly due to a pressure caused by friction generated while extending a hair shaft on a solid surface or in the absence of sebum (Sundberg et al, 2000) . Although not proven, we speculate that the critical modification was our method of immobilizing the hair shaft in silicon grease that provided friction-free environment; anchoring the tip of shafts may have allowed gravity to facilitate the separation of club hairs from the hair matrix and that supports new hair shaft formation. In addition, the smaller size of mouse vibrissae, in comparison with human or rat vibrissae, could also be a contributing factor in the anagen reinitiation in vitro As this culture system is the first to support reinitiation of anagen in culture dish, we hope this improvement will expedite the testing and development of hair cycle modifying pharmaceuticals.