Cell Biochemistry and Biophysics

A number of experimental and mathematical problems must be solved before high resolution physical maps of mammalian chromosomes can be reliably determined. Such a map might consist of an ordered set of nonsequenced, overlapping DNA fragments 20,000-40,000 bases long, produced by digestion of a chromosome, using two restriction enzymes. Map construction requires assigning a signature to each fragment that differentiates it unambiguously from every other fragment, and then devising a computationally efficient algorithm that will provide a unique ordering of the fragments. In the first part of this paper we present a polynomial time algorithm that yields a unique map, and is largely independent of the method for assigning signatures. In the next section we analyze the distribution of lengths of restriction digest fragments and discuss the implications for the algorithm, including the expected number of map gaps. Finally, we discuss a specific method for assigning signatures proposed by Hans Lehrach, based on which of a panel of probes binds to a given fragment. In particular we examine the effects of fragment length heterogeneity on the theoretical optimum length and number of probes, and the extent to which false signatures might be obtained by nonspecific binding. We conclude that the Lehrach strategy is effective provided the number of probes is >-150, but that each fragment will need testing with at most 25 probes.

The objective of this study was to analyze the clinical manifestation, imaging characteristics, genotype, and the relationship between the three aforementioned parameters in two pedigrees suffering from spinocerebellar ataxia. To evaluate the clinical manifestation of the two pedigrees and to compare the characteristics, we performed the MRI analysis of some patients from both pedigrees, while 2 ml of the peripheral blood sample was collected for gene analysis. The gene analysis data showed that pedigree 1 was certified spinocerebellar ataxia type-2 (SCA2); the CAG repeats in the proband, proband’s mother, and proband’s brother were 44, 36, and 38, respectively. The MRI revealed brainstem cerebellar atrophy and “cross sign” and “ordinate sign” of pons. Pedigree 2 was certified SCA1; the CAG repeats of the proband, proband’s aunt, and proband’s asymptomatic cousin were 60, 51, and 52, respectively. The MRI revealed cerebellar atrophy in these individuals. We, therefore, concluded that it was difficult to diagnose the SCA subset solely through the clinical manifestation. The imaging characteristics analysis and final diagnosis depended basically on gene analysis data.

Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is useful for lung cancer diagnosis and lymph node staging. The purpose of this study was to investigate EBUS-TBNA for managing mediastinal and hilar lymphadenopathies without intrapulmonary masses. We retrospectively reviewed our EBUS-TBNA database that was obtained between August 2010 and October 2012. Mediastinal and hilar lymphadenopathies of unknown origin and in the absence of known pulmonary malignancies were included. Final diagnoses were determined by EBUS-TBNA, surgery, and/or clinical follow-up for at least 6 months. Sensitivity, specificity, accuracy, and positive and negative predictive values were determined using standard statistical methods. We identified 128 patients with mediastinal and hilar lymphadenopathies and without intrapulmonary masses. EBUS-TBNA was successfully performed to obtain samples from 161 lymph nodes and mediastinal masses. EBUS-TBNA was diagnostic for 119 of 128 patients (93.0 %) for all disease categories. The sensitivity, specificity, positive predictive value, negative predictive value, and overall accuracy of EBUS-TBNA were 89.8, 100, 100, 81.6, and 93.0 %, respectively. The procedures were uneventful and there were no severe complications. EBUS-TBNA is a safe, minimally invasive approach for diagnosing mediastinal and hilar lymphadenopathies without intrapulmonary masses. It obviates the need for more invasive procedures for tissue sampling of the mediastinum and hilum.

The aim of this study was to assess myocardial dysfunction in primary diabetes patients with microalbuminuria by 2-dimensional speckle tracking strain. Sixty-two patients with diabetes with or without hypertension and 37 matched hypertension controls were consecutively recruited from January 2011 to 2013. Routine physical examinations, laboratory tests, and echocardiography were performed in all patients. Subjects enrolled were divided into three groups according to history and urine albumin/creatinine ratio (ACR): group I: patients with only hypertension and normoalbuminuria (ACR < 30 mg/g), group II: patients with both hypertension and diabetes and normoalbuminuria (ACR < 30 mg/g), and group III: patients with both hypertension and diabetes and microalbuminuria (ACR 30–300 mg/g). Echocardiographic images of three cardiac cycles were acquired for off-line analysis using the GE EchoPAC software. Indices of cardiac function, including longitudinal, radial and circumferential strains, torsion, and left ventricular ejection fraction (LVEF) were assessed. Statistical analysis was performed using SPSS 13.0. Finally, 56 subjects and 32 controls were included in the analyses. There was no significant difference in age, gender, heart rate, BMI, and LVEF among groups, except for the blood pressure, ACR, and HbA1c. E wave, A wave, EDT, E m, and E/E m in group III were different with those in group I. Mean longitudinal strain (mSL), average SL of six segments in 4-chamber apical view (SL4) decreased obviously. The peak circumferential strain decreased in group III, while the torsion was compensatively increased. ACR was negatively related to mSL, SL4, E/E m, and positively related to torsion. We deduced that ACR maybe a predictor for myocardial damage in primary diabetes.

Hypertension leads to impaired contractile function. This study examined the impact of inhibition of sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) by thapsigargin or cyclopiazonic acid (CPA) on cardiac contractile function in ventricular myocytes from Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR). Mechanical properties were examined including peak shortening (PS), time-to-PS (TPS), time-to-90% relengthening (TR90), and maximal velocity of shortening/relengthening (±dL/dt). Intracellular Ca2+ transients were evaluated as fura-2 fluorescent intensity (FFI), excitation-induced change in FFI (ΔFFI=peak-basal), and fluorescence decay rate (τ). Expression of Ca2+ regulatory proteins SERCA2a, Na+−Ca2+ exchanger (NCX), and phospholamban (PLB) were assessed by reverse transcriptase polymerase chain reaction and Western blot. SHR rats exhibited elevated blood pressure. SHR myocytes displayed decreased PS±dL/dt, peak FFI, and ΔFFI; shortened TPS; prolonged τ with normal TR90; and basal FFI compared with WKY myocytes. Inhibition of SERCA with thapsigargin (5μM) or CPA (10 μM) significantly depressed PS±dL/dt, baseline FFI, and ΔFFI, and prolonged TPS, TR90, and τ in WKY myocytes. However, SHR myocytes were relatively insensitive to thapsigargin or CPA with only TPS and TR90 prolonged. Both mRNA and protein expressions of NCX and PLB were significantly enhanced, whereas SERCA2a protein abundance was reduced in SHR rats compared with the WKY group. Our data suggest that inhibition of SERCA function differentially affected cardiac contractile function in ventricular myocytes from normotensive and hypertensive rats possibly through reduced SERCA2a, elevated PLB, and NCX expression under hypertension.

Previous studies have shown thatcis unsaturated free fatty acids (uFFAs) are able to cause alterations in the normal distribution pattern of certain cytoskeletal proteins in lymphocytes, including tubulin, actin, α-actinin, and myosin. The cytoskeletal protein spectrin naturally possesses a marked heterogeneity of distribution among resting T and B lymphocytes isolated from all murine lymphoid organs. In some cells, spectrin is observed in a ring-like staining pattern at the periphery of the cell, reflecting a likely association with the cell membrane; in other cells, spectrin is found within the cytoplasm as a large single aggregate or in several smaller aggregates. Addition of uFFA to freshly isolated murine lymphocytes causes disruption in the latter pattern of spectrin organization. Following short-term incubation (15 min) of tissue-derived lymphocytes (from spleen, thymus, and lymph node) and 1 μg/mL uFFA (oleic [18:1cis], linoleic [18:2cis, cis], arachidonic [20:4], or elaidic [18:1trans] acid) there is a loss of cytoplasmic aggregates of spectrin and a concomitant increase in cells in which spectrin is diffusely distributed. This effect is not seen when two saturated FFAs (sFFAs) were used. When using DO11.10 cells, a T-cell hybridoma in which nearly all cells constitutively express a cytoplasmic aggregate of spectrin, a similar effect was observed, but greater concentrations (10–20 μg/mL) of FFA were needed to obtain the same effect. Addition of calcium to the incubation buffer substantially blocks spectrin reorganization. In several disease states, serum levels of FFA are observed to be excessively high; our data support the hypothesis that cytoskeletal reorganization in lymphocytes may be related to the altered immune function frequently observed in these conditions.

Malignant obstructive jaundice is caused by tumors arising from the head of the pancreas and biliary tree, or seen due to secondary metastases in the porta hepatis lymph nodes. Magnetic resonance cholangiopancreatography (MRCP) is a non-invasive diagnostic technique that can be used for imaging the entire biliary tree and pancreatic duct system. The objective of this study was to evaluate the accuracy of MRCP in the diagnosis of malignant obstructive jaundice. The methods used involved comparative review of the images obtained by using magnetic resonance imaging and MRCP as well as comparison between MRCP- and pathology-based diagnoses. The accuracy of MRCP-based diagnosis of malignant obstructive jaundice was analyzed. Our data show that the positive rate of anatomical diagnosis and the detection rate of bile ducts on the proximal side of obstruction are 100%. The diagnostic accuracy of malignant obstruction was 82.9%. MRCP was found to have high diagnostic specificity for determining the location and extent of obstruction. We, therefore, concluded that MRCP had significance for clinical diagnosis of malignant obstructive jaundice. The positive rate of localization diagnosis was 100%. Distinguishing the quality of obstruction was also important. The diagnostic accuracy of MRCP for malignant obstructive jaundice was remarkably higher.

Ferroptosis is a newfound mode of regulated cell death that may have potential to associate with prognostic or diagnostic factors in glioma. In this research, 5 genes related to glioma were screened through the FerrDb database, and we analyzed the combination between genes and glioma of survival and prognosis via TCGA, GEPIA, TIMER, and other databases. Survival curve and prognostic analysis showed that the overexpression of NFE2L2 and NOX4, respectively, has a remarkable link with a worse prognosis in glioma. Then, the association between the expression of the two genes and tumor-infiltrating immune cells level was explored based on the GSCA, and the immunity of NFE2L2 and NOX4 based on the TISIDB database was also investigated. In glioma, especially GBM, there is a strong association between gene expression and immune infiltration, even in macrophages, nTreg, and Th2 cells, which play immunosuppressive functions in TME. In conclusion, these results indicate that NFE2L2 and NOX4 could be risk prognosis biomarkers in glioma, and they bound up with immune infiltration and tumor immunity in tumorigenesis.

The objective of this study is to explore the relationship between the hypersensitive c-reactive protein (hs-CRP) level and the prognosis of acute brainstem infarction. Serum levels of hs-CRP were measured in 68 patients with acute brainstem infarction 72 h after disease onset. The hs-CRP levels in the U.S. National Institutes of Health Stroke Scale (NIHSS) score group and in the modified RANKIN scale (mRS) score group were compared. The independent risk factors of brainstem infarction were analyzed using Logistic binary regression. The hs-CRP level was significantly higher in the group with NIHSS >5 compared with the one with NIHSS ≤5 (P = 0.004). In the group with mRS > 2, the age, smoking history, and blood glucose level were significantly higher than those in the group with mRS ≤ 2 (P < 0.05), whereas the hs-CRP level was significantly higher (P = 0.001). Age and hs-CRP level were the independent prognostic factors of the brainstem infarction. The serum hs-CRP level is closely related with the severity and prognosis of brainstem infarction, and is an independent risk factor of acute brainstem infarction.