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Background: Diabetic retinopathy (DR) is a specific neurovascular complication of diabetes mellitus (DM). Clinically, family history is a widely recognized risk factor for DR, assisting diagnosis and risk strata. However, among a great amount of DR patients without hereditary history like hypertension and diabetes, direct and simple risk factors to assist clinical decisions are still required. Herein, we intend to investigate the associated risk factors for these DR patients based on systemic inflammatory response indexes, neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR)., Methods: We consecutively enrolled 1030 patients with a definite diagnosis of type 2 diabetes mellitus (T2DM) from the endocrinology department of the Second hospital of People in Yun Nan. Based on funduscopy and family history checking, we excluded patients with a family history of hypertension and diabetes and finally enrolled 264 patients with DR and 206 patients with non-diabetic retinopathy (NDR). Through correlation analysis, univariate and multivariate regression, we further explore the association between NLR, PLR, and DR. On top of that, we investigate the effect of NLR and PLR on risk reclassification of DR., Results: Compared with NDR patients, NLR and PLR levels are significantly higher among DR patients (NLR: 2.36 ± 1.16 in DR group versus 1.97 ± 1.06 in NDR group, p < 0.001; PLR: 11.62 ± 4.55 in DR group versus10.56 ± 4.45 in NDR group, p = 0.012). According to univariate analysis, NLR and PLR add risks to DR. After fully adjusting co-founders, NLR, as both continuous and categorical variate, remains an independent risk factor for DR (OR (95%CI): 1.37 (1.06, 1.78) P = 0.018). And though PLR was not independently associated with DR as a continuous variable (OR (95%CI) 1.05 (0.99, 1.11) p = 0.135), the highest quantile of PLR add two-fold increased risk (OR (95%CI) 2.20 (1.05, 4.59) p = 0.037) in the fully adjusted model for DR. In addition, addition of PLR and NLR to the established factor hemoglobin (Hb) improved the discriminability of the model and assisted the reclassification of DR. After combining PLR and NLR the Area under curve (AUC) of Hb based model raised from 0.76 to 0.78, with a category-free net reclassification improvement (NRI) of 0.532 (p < 0.001) and integrated discrimination improvement (IDI) of 0.029 (p < 0.001)., Conclusions: Systemic inflammatory response indexes NLR and PLR were associated with the presence of DR among patients without associated family history and contributed to improvements in reclassification of DR in addition to Hb., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s) 2020.)

Objective: To explore the clinical features and risk factors of hepatic injury due to immune checkpoint inhibitors (CPI) therapy in malignant tumor. Methods: Data of 112 patients (64 men and 48 women) who received CPI between January 2016 and March 2019 in Chinese Academy of Medical Sciences and Peking Union Medical College Shenzhen Hospital, and Huazhong University of Science and Techology Union Shenzhen Hospital were retrospectively collected. The median age of these patients was 60 years. Results: Hepatic adverse events were observed in 30 patients out of 112 patients (26.8%). Among them, the incidence of grade 3-5 hepatic adverse events were 7.14% (8/112). The median time of hepatic adverse event occurrence was 3 weeks (2-30) after undergoing therapy. The results of univariate and multivariate analyses showed that liver cancer was attributed to the CPI induced hepatitis ( P <0.05). Patients with severe hepatic injury got almost complete resolution after receiving methlprednisolone for 4 to 6 weeks. Conclusion: Live cancer is the risk factor of CPI-related hepatic adverse events.

In this paper, we develop a novel subspace-based recovery algorithm for non-blind deconvolution (named SND). With considering visual importance difference between image structures and smoothing areas, we propose subspace data fidelity for protecting image structures and suppressing both noise and artifacts. Meanwhile, with exploiting the difference of subspace priors, we put forward differentiation modelings on different subspace priors for improving deblurring performance. Then we utilize the least square integration method to fuse deblurred estimations and to compensate information loss of subspace deblurrings. In addition, we derive an efficient optimization scheme for addressing the proposed objective function by employing the methods of least square and fast Fourier transform. Final experimental results demonstrate that the proposed method outperforms several classical and state-of-the-art algorithms in both subjective and objective assessments.

In the natural environment, arsenic (As) is mainly adsorbed on iron oxide minerals. The release of adsorbed arsenic from iron oxide minerals to the water is the main source of arsenic pollution. Microbes play a crucial role for this process. The purpose of this study was to investigate the effect of the sulfate-reducing bacteria Desulfovibrio vulgaris DP4 on the transformation and mobilization of As. The experimental results show that the released As concentration of the two systems is 0 μmol·L -1 at 0 h. Compared with the control, DP4 promotes the desorption of As(Ⅴ) before the 84 h incubation process. The released As concentration reaches the maximum value of 12.6 μmol·L -1 at 13 h, accounting for~79% of the initial total adsorbed As (16 μmol·L -1 ). The maximum released As concentration is~8.4 times higher than that of the control (1.5 μmol·L -1 ). After 84 hours, the concentration of the released As in the DP4 system is lower than the abiotic control, which suggests that the released As is readsorbed on the solid surface. During the incubation process, the As mobility is significantly correlated with Eh. The XRD results show that the crystallinity of the solid samples in the DP4 system decreases by~50%. In general, a lower crystallinity of the adsorbent indicates a higher adsorption capacity. This may be one important reason for the As readsorption after 84 h. In addition, the SEM shows that goethite is agglomerated by DP4 and the EDS results indicate that goethite is partially transformed to an Fe-S mineral. Based on XANES, arsenic-sulfur minerals were not detected in the solid phase, which further confirms the SEM-EDS results, that is, that Fe-S minerals formed in the solid phase, rather than As 2 S 3 (AsS). The released As was readsorbed on the secondary iron mineral, resulting in a lower dissolved As concentration in the DP4 system than in the abiotic control. Furthermore, 19% As(Ⅲ) was detected in the solid phase while dissolved As(Ⅲ) was not determined during the incubation process. The results suggest that sulfate-reducing bacteria may directly reduce adsorbed As(Ⅴ) to As(Ⅲ).

To improve the coagulation performances, new composite coagulants were used to treat different water samples. The results indicate that Ca 2+ has no significant effects on the removal efficiency for turbidity in the kaolin system. The residual aluminum decreased from 0.15 mg·L -1 to 0.10 mg·L -1 (AlCl 3 was used as coagulant and the coagulant dosage was 0.10 mmol·L -1 ). The presence of Ca 2+ led to the decrease of the amount of negative charges in the HA system and the residual aluminum decreased due to the decrease of the complexation between the HA molecules and Al-based coagulants. When the raw water contained BSA molecules and the coagulant dosage was 0.16 mmol·L -1 , the flocs formed by PACl with Ca 2+ were larger (~50 μm) than the flocs generated by PACl and the settleability also improved. Under alkaline conditions (pH=8.5), the DOC concentration decreased after coagulation process by~0.2-0.6 mg·L -1 and the residual aluminum decreased by~0.4-0.7 mg·L -1 using composite coagulants. Under acidic conditions (pH=5.5), the concentrations of DOC and residual aluminum did not significantly differ.

Bubble-driven micromotors have attracted substantial interest due to their remarkable self-motile and cargo-delivering abilities in biomedical or environmental applications. Here, we developed a hollow micromotor that experiences fast self-propulsion underneath an air-liquid interface by periodic bubble growth and collapse. The collapsing of a single microbubble induces a ∼1 m·s -1 impulsive jetting flow that instantaneously pushes the micromotor forward. Unlike previously reported micromotors propelled by the recoiling of bubbles, cavitation-induced jetting further utilizes the energy stored in the bubble to propel the micromotor and thus enhances the energy conversion efficiency by 3 orders of magnitude. Four different modes of propulsion are, for the first time, identified by quantifying the dependence of propulsion strength on microbubble size. Meanwhile, the vertical component of the jetting flow counteracts the buoyancy of the micromotor-bubble dimer and facilitates counterintuitive hovering underneath the air-liquid interface. This work not only enriches the understanding of the propulsion mechanism of bubble-driven micromotors but also gives insight into the physical aspects of cavitation bubble dynamics near the air-liquid interface on the microscale.

In order to ensure drinking water quality, three different Al-based coagulants [Al 2 (SO 4 ) 3 (AS), Al 13 , Al 30 ] were used to treat water laden with different algae [ Microcystis aeruginosa (cyanobacteria), Chlorella (green algae), Cyclotella (diatoms)]. Floc size, strength factor, and recovery factor under different conditions were measured to investigate the mechanisms in the coagulation-ultrafiltration process. The results indicated that the main mechanism in the coagulation process using Al 13 or Al 30 as coagulants was electrostatic patching and the main mechanism using AS was charge neutralization. In the Microcystis aeruginosa and Chlorella systems, when Al 13 and Al 30 were used as coagulants at low dosage, the coagulants that were adsorbed on the side (which existed on the surfaces of Microcystis aeruginosa and Chlorella cells) would lose their ability to aggregate the algal cells. When AS was used as coagulant, the electric double layer was effectively compressed. The repulsive force between algal particles decreased, and the flocs formed easily. In the Cyclotella system, Al 13 and Al 30 coagulants effectively formed the flocs through electrostatic patch effects. There was a significant correlation between membrane flux and floc size, and the larger flocs formed a looser cake layer on the membrane surface.

Stem cells from human exfoliated deciduous teeth (SHEDs) have great potential to treat various dental-related diseases in regenerative medicine. They are usually maintained with 10% fetal bovine serum (FBS) in vitro. Modified platelet-rich plasma (mPRP) would be a safe alternative to 10% FBS during SHEDs culture. Therefore, our study aimed to compare the proliferation and differentiation of SHEDs cultured in mPRP and FBS medium to explore an optimal concentration of mPRP for SHEDs maintenance. Platelets were harvested by automatic blood cell analyzer and activated by repeated liquid nitrogen freezing and thawing. The platelet-related cytokines were examined and analyzed by ELISA. SHEDs were extracted and cultured with different concentrations of mPRP or 10% FBS medium. Alkaline phosphatase (ALP) activity was measured. Mineralization factors, RUNX2 and OCN, were measured by real-time PCR. SHEDs were characterized with mesenchymal stem cells (MSCs) markers including vimentin, CD44, and CD105. mPRP at different concentrations (2, 5, 10, and 20%) enhanced the growth of SHEDs. Moreover, mPRP significantly stimulated ALP activity and promoted expression of RUNX2 and OCN compared with 10% FBS. mPRP could efficiently facilitate proliferation and differentiation of SHEDs, and 2% mPRP would be an optimal substitute for 10% FBS during SHEDs expansion and differentiation in clinical scale manufacturing.

The As removal process and mechanism from groundwater using ferric flocculation-direct filtration system was investigated using batch, field pilot tests, extended X-ray absorption fine structure ( EXAFS) spectroscopy, and charge-distribution multisite complexation (CD-MUSIC) model. The results showed that arsenate [As(V)] was the dominant As species in the groundwater with a concentration of 40 μg x L(-1). The treatment system could supply 64 984 L As-safe drinking water (< 10 μg L(-1)) using Fe 1.5 mg x L(-1). Toxicity characteristic leaching procedure (TCLP) demonstrated that the leachate As was 3.4 μg x L(-1), much lower than the EPA regulatory concentration (5 mg x L(-1)). EXAFS and CD-MUSIC model indicated that As(V) was adsorbed onto ferric hydroxide via bidentate binuclear complexes in the pH range of 3 to 9.5, while formation of precipitate with Ca or Mg dominated the As removal at pH > 9.5.

Objective: To study the therapeutic effect and safety of Longjintonglin Capsule in the treatment of type III prostatitis (chronic prostatitis/chronic pelvic pain syndrome, CP/CPPS)., Method: We selected 240 patients with type III prostatitis according to the diagnostic standards of the American National Institute of Health (NIH) and treated them with Longjintonglin Capsule orally 3 capsules once tid for 12 weeks. Based on the NIH chronic prostatitis symptom index (NIH-CPSI), traditional Chinese medicine (TCM) syndrome score, and leukocyte count in the expressed prostatic secretion (EPS), we evaluated the results of treatment., Results: Totally 238 patients completed the treatment, including 108 IIIA and 120 III B prostatitis cases. Before and after 4, 8, and 12 weeks of treatment, the total NIH-CPSI scores were 23.12 ± 6.99, 18.22 ± 6.39, 14.12 ± 5.88, and 12.36 ± 6.04 (P < 0.01) in the IIIA prostatitis patients and 22.01 ± 6.28, 17.56 ± 5.89, 13.67 ± 5.18, and 11.45 ± 5.22 in the III prostatitis patients (P < 0.01), the TCM syndrome scores were 52.12 ± 13.08, 48.13 ± 12.11, 43.05 ± 11.19, and 40.78 ± 10. 59 in the former (P < 0.01) and 53.02 ± 12.12, 49.32 ± 12.78, 44.01 ± 11.79, and 39.67 ± 10.26 in the latter (P < 0.01), and the leukocyte counts were 26.09 ± 21.55, 23.02 ± 18.61, 18.25 ± 17.79, and 15.36 ± 16.38 in the IIIA cases (P < 0.01). Neither abnormalities in liver and renal function nor obvious adverse events were observed during the experiment., Conclusion: Longjintonglin Capsule, with its advantages of safety, effectiveness, and no obvious adverse reactions in the treatment of type III prostatitis, deserves to be recommended for clinical application.

Purpose: This study used spiral computed tomography to identify the anterior loop of the inferior alveolar nerve, and to measure its length and position in Chinese. This information may be useful to safely install endosseous implants in the most distal area of the interforaminal region., Methods: Sixty-eight Chinese patients were included in this retrospective study. Patients were scanned by 64-slice spiral computed tomography, and the prevalence, length, and position of the anterior loop were assessed using the multiplanar capabilities of software., Results: An anterior loop could be identified in 83.1% of the cases, with a mean length of 2.09 mm (range 0-5.31 mm). The mean distance from the superior border of the mental foramen to the alveolar crest was 13.00 mm, and the mean distance from the superior border of the origin of the anterior loop to the alveolar crest was 17.83 mm., Conclusions: The anterior loop was highly prevalent in Chinese, and the length of the anterior loop was highly variable. Therefore, we recommend that drilling commences from a location approximately 5.5 mm mesially from the mental foramen, when installing implants in the most distal interforaminal area.

In order to enhance the hydrophilicity of the membrane and improve the antifouling properties, poly (2-hydroxyethyl methacry-late) (poly(HEMA)) was grafted to the surface of the poly(vinylidene fluoride) (PVDF) membrane by using the atom transfer radical polymerization (ATRP) method. After that, ceric ammonium nitrate (CAN) as the initiator, N, N'-methylene bisacrylamide as a cross-linking agent, a zwitterionic polymer, poly (3-( methacryloylamino) propyl-dimethyl-(3-sulfopropyl) ammonium hydroxide) (poly(MPDSAH)) were successfully grafted onto the membrane surface by radical polymerization reaction. Attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy, X-ray photoelectron spectroscopy (XPS), scanning electron microscope (SEM) and contact angle measuring were employed to analyze the property and the morphology of structures before and after the membrane surface-modification. The grafting density (GD) gradually increased, with the grafting time increasing, the pore size of the membrane became smaller, and the porosity decreased, but the surface hydrophilicity of membrane was significantly enhanced at the same time. The adsorption of bovine serum albumin (BSA) tests and filtration experiments were carried out to investigate anti-fouling performances of membrane before and after modification. With the GD increasing, the amount of adsorption on the film surface significantly reduced in the high-concentration BSA solution. The water contact angle (CA) decreased most, from 77. 2 degrees to 41.7 degrees within 5 s to 0, and a flux recovery ratio up to 94. 961% , when the GD reached 288. 340 microg.cm-2. Therefore, the optimal grafting time was 2 h, with the grafting density of 288. 340 microg.cm-2.

Paulownia kawakamii is a fast-growing timber tree. In this study, 21 primer sets were developed using an enriched genomic library. The genetic diversity was measured in one P. kawakamii population. The number of alleles per locus ranged from 2 to 19. The observed and expected heterozygosities varied from 0.158 to 0.842 (mean = 0.421) and from 0.376 to 0.952 (mean = 0.771), respectively. All 21 loci were also polymorphic in closely related species (P. tomentosa, P. elongata, and P. fortunei). The described markers will be useful in future population genetic studies and molecular breeding of these Paulownia species.

Semen-derived enhancer of viral infection(SEVI) is a peptide fragment (PAP248-286) from prostatic acid phosphatase(PAP), which can enhance human immunodeficiency virus infection. The mechanisms of SEVI include: (1) SEVI with several cationic amino acid residues reduced electrostatic repulsion between HIV virus and the target cells; (2) The disorder state of SEVI in the human body fluids was helpful to the interaction between virus and the target cell membranes; (3) SEVI could capture HIV particles directly and speed the velocity of virus on the surface of the target cells and improve adsorption and fusion. Currently, the substances of inhibiting SEVI activity include: EGCG from green tea, small molecule compound of aminoquinoline Surfen, ThT analogs BTA-EG6. Those compounds might block the combination of HIV and SEVI or prevent the formation of amyloid fibers, and then reduce the enhancement of SEVI. The studies on the biological characteristics and mechanisms of SEVI have a big benefit for the prevention and treatment of HIV infection.

Objective: To investigate the antiviral activity of 3-hydroxyphthalic anhydride-modified ovalbumin (HP-OVA) against herpes simplex virus 2 (HSV-2) in vitro., Methods: By chemical modification, ovalbumin (OVA) was treated with 3-hydroxyphthalic anhydride (HP) to prepare HP-OVA. The anti-HSV-2 activity against HSV-2 333 virus in vitro and the cytotoxicity of HP-OVA in African green monkey kidney cells (Vero cells) were detected by MTT colorimetric assay. The inhibitory effects of HP-OVA on 17 strains of vaginal lactobacilli were observed by microscopy., Results: Anhydride-modified ovalbumin significantly inhibited the infection by HSV-2 with an IC(50) of 23.56±8.33 µg/ml. HP-OVA showed only low cytotoxicity to the host cells with a CC(50) over 1 mg/ml. HP-OVA did not produce significant inhibitory effect on the 17 strains of vaginal lactobacilli (MIC>1 mg/ml)., Conclusion: Anhydride-modified protein HP-OVA exhibits potent anti-HSV-2 activity in vitro and can be a good microbicide candidate for prevention of sexually transmitted diseases.

Objective: To explore the effectiveness of voice training on professional voice users with voice disorders., Methods: Thirty one professional voice users suffering from voice symptoms without organic disorder were treated with a four-week voice training program, including vocal hygiene education, respiratory training and resonant therapy. Voice handicap index, maximum phonation time, noise to harmonic ratio, jitter and shimmer were compared before and after the training., Results: The voice handicap index (x(-) ± s) decreased from 33.7 ± 19.2 to 18.8 ± 18.4 after the training and the difference was significant (t = 6.14, P < 0.05). Maximum phonation time (x(-) ± s) increased significantly from (15.5 ± 5.8) s to (18.6 ± 6.0) s (t = -3.43, P < 0.05). jitter (M [P(25); P(75)]) decreased significantly after the training from 0.42% [0.36%; 0.62%] to 0.35% [0.29%; 0.47%] (Z = -2.51, P < 0.05), while there was no significant change in shimmer and noise to harmonic ratio., Conclusion: Voice training can effectively reduce the vocal symptoms in professional voice users, decrease the handicap degree and increase the respiratory support during phonation, thus improve the voice quality.

Objective: To investigate the association of recurrent aphthous ulcer (RAU) with Helicobacter pylori (Hp) infection and digestive diseases., Methods: Saliva samples were collected from 82 patients with RAU and 74 healthy volunteers for Hp detection with PCR., Results: The positivity rates of HP differed significantly between RAU patients and healthy volunteers (43.9% vs 16.2%, P<0.001). In the 82 RAU patients, 22 (26.82%) were identified to have gastritis and peptic ulcer, whereas only 7 out of the 74 healthy volunteers (10.45%) had such digestive diseases, showing significant difference between them (P<0.01)., Conclusion: Hp might in some way associate with RAU, which in turn is associated with an increased incidence of digestive diseases.

We compared the urothelial injury to the bladder caused by four agents capable of dissolving calcium salts. The solutions were administrated in an antegrade way through left ureterostomies in 54 rabbits for periods of 24, 48 and 72 h. The bladders were then removed and three routine histological sections were made for each. The following six solutions were used: physiological sodium chloride solution (Phys), artificial urine (Art), 0.03 M disodium EDTA buffered to pH 8.5 with triethanolamine (EDTA), 10% Renacidin (R), test solution 2 (S2, using D-gluconic acid-lactone and other compounds that differ from R in terms of ingredients or quantity), and test solution 1 (S1, using D-gluconic-acid instead of D-gluconic acid-lactone in S2 but keeping the other ingredients the same) for irrigation. At 24 h there was no observable urothelial damage caused by perfusion with Phys or Art; solutions R, S1 and S2 caused approximately the same level of injury to the rabbit bladder mucosa; however, irrigation with disodium-EDTA caused more serious urothelial injury than R, S1 and S2 (P<0.05, chi2-test) and may be unacceptable. The damage to bladder tissues treated with S1 and S2 was less than that caused by R, but this was not significant (P>0.05, chi2-test). Following a prolonged irrigation time, all of these solutions cause further urothelial damage, but EDTA caused the most, followed by R, S1, S2, Phys or Art, respectively, at 48 and 72 h. In view of the better solubility effect of solutions S1 and S2 compared with R, it might be justified in accepting the more pronounced urothelial irritation caused these solutions, but in order to enhance their effectiveness and reduce urothelial injury further study will be needed.

Objective: To evaluate the effectiveness of free graft transplantation two-stage urethroplasty for hypospadias repair., Methods: Fifty-eight cases with different types of hypospadias including 10 subcoronal, 36 penile shaft, 9 scrotal, and 3 perineal were treated with free full-thickness skin graft or (and) buccal mucosal graft transplantation two-stage urethroplasty. Of 58 cases, 45 were new cases, 13 had history of previous failed surgeries. Operative procedure included two stages: the first stage is to correct penile curvature (chordee), prepare transplanting bed, harvest and prepare full-thickness skin graft, buccal mucosal graft, and perform graft transplantation. The second stage is to complete urethroplasty and glanuloplasty., Results: After the first stage operation, 56 of 58 cases (96.6%) were successful with grafts healing well, another 2 foreskin grafts got gangrened. After the second stage operation on 56 cases, 5 cases failed with newly formed urethras opened due to infection, 8 cases had fistulas, 43 (76.8%) cases healed well., Conclusions: Free graft transplantation two-stage urethroplasty for hypospadias repair is a kind of effective treatment with broad indication, comparatively high success rate, less complications and good cosmatic results, indicative of various types of hypospadias repair.

Background: Acute renal allograft rejection associated with the development of donor-specific alloantibody (acute humoral rejection, AHR) typically carries a poor prognosis. The best treatment of this condition remains undefined., Methods: During a 14-month period, 73 renal transplants were performed. During the first postoperative month, five recipients (6.8%) with AHR were identified. The diagnosis was based on: (1) evidence of severe rejection, resistant to steroid and antilymphocyte therapy; (2) typical pathologic features; and (3) demonstration of donor-specific alloantibody (DSA) in recipient's serum at the time of rejection. Pretransplant donor-specific T- and B-cell cross-matches were negative., Results: Plasma exchange (PE, four to seven treatments per patient) significantly decreased circulating DSA to almost pretransplant levels in four of five patients, and improvement in renal function occurred in all patients. One patient had recurrent renal dysfunction in the setting of an increase in circulating DSA. A second series of five PE treatments decreased DSA and reversed the rejection episode. Rescue therapy with tacrolimus (initial mean dose: 0.14+/-0.32 mg/kg/day) and mycophenolate mofetil (2 g/day) was used in five of five and four of five patients, respectively. With a mean follow-up of 19.6+/-5.6 months, patient and allograft survival are 100%. Renal function remains excellent with a mean current serum creatinine of 1.2+/-0.3 mg/dl. (range: 0.9-1.8 mg/dl)., Conclusions: Our findings suggest that a therapeutic approach combining PE and tacrolimus-mycophenolate mofetil rescue has the potential to improve the outcome of AHR.

CD4+ T cells play a crucial role in the development of lupus in MRL-lpr/lpr mice: incomplete deletion/silencing of self-reactive CD4+ T cells leads to T cell activation, which causes both polyclonal B cell activation and T cell infiltration of multiple organs. Furthermore, anti-CD4 antibody therapy ameliorates disease and prolongs survival. Because CD4 is normally involved in both tolerance induction and T cell activation, we questioned whether signaling through CD4 was normal among T cells in this strain. For this purpose, signal transduction in CD4+ T cells derived from MRL-lpr/lpr and normal mice were compared, using an autoreactive CD4+ T cell clone and freshly isolated CD4+ T cells derived from mice of varying ages. Tyrosine phosphorylation was similar among MRL and normal CD4+ T cells after cross-linking with either anti-TCR antibody or anti-CD3 antibody, and following co-culture with Con A. In constrast, cross-linking of surface CD4 resulted in deficient tyrosine phosphorylation of cellular proteins in MRL T cells. By comparison, lck protein expression in MRL CD4+ T cells was found to be lower than normal. However, following stimulation with Con A, lck enzyme activity, as detected by autophosphorylation of lck, was comparable in MRL and normal T cells. The observed differences were present in the autoreactive T cell clone as well as in T cells isolated from both pre-diseased and diseased mice, and they could not be explained by variation in surface density of CD4. These results raise the possibility that abnormal signaling through CD4 may contribute to impaired tolerance and expansion of autoreactive T cells exhibited in MRL-lpr/lpr mice.

The direct effects of varying concentrations (5-40 mM) of D,L-carnitine were studied in two populations, subsarcolemmal and interfibrillar, of cardiac mitochondria exposed to inorganic phosphate (Pi). After 5 min preincubation 20 mM Pi significantly depressed oxidative phosphorylation rate and ADP/ATP translocase activity, in both populations. Inclusion of D,L-carnitine during preincubation significantly prevented the Pi-induced depression in oxidative phosphorylation without affecting the ADP/ATP translocate system. The Pi-induced inhibition in mitochondrial oxygen consumption rate was seen with either pyruvate-malate, glutamate-malate or succinate as respiratory substrates and was also observed in uncoupled mitochondria treated with 2,4-dinitrophenol. Mitochondrial swelling and shrinkage studies revealed Pi-induced inner membrane instability, a phenomenon prevented by D,L-carnitine in a dose-dependent manner. The effect of Pi was also observed at a concentration of 5 mM which was also prevented by carnitine. Mepacrine, a phospholipase A2 inhibitor, failed to prevent any of the effects of Pi. The results therefore suggest that Pi can produce a depression in mitochondrial oxidative phosphorylation through a mechanism possibly associated with disturbed inner membrane structure and function but apparently unrelated to phospholipase A2 activation. The salutary actions of carnitine may partly explain its protective effects in the ischemic and reperfused heart, a phenomenon associated with enhanced intracellular Pi accumulation.

The effects of exogenous phosphatidylcholine (PC) on some potential mechanisms of ischemia and reperfusion-induced arrhythmias were tested using the superfused right ventricular free wall of the guinea pig. Exposure of the preparation to simulated "ischemia" (hypoxia, acidosis, glucose deprivation and hyperkalemia) resulted in several electrophysiological derangements, including a marked depolarization of the maximum diastolic potential (MDP) in both endocardium and epicardium, shortening of the action potential duration (APD), and prolongation of the transmural conduction time followed by transmural conduction block. In a few preparations, coupled beats were also observed. Reperfusion was associated with arrhythmic activity in all preparations. Both the characteristics and the severity of reperfusion-associated arrhythmias were dependent upon the duration of the preceding "ischemia". In hearts exposed to ischemic conditions for 40 min, transmural conduction block persisted until 45 min of reperfusion and no electrical activity was present in the epicardium during this time. However, both coupled beats as well as abnormal automaticity were observed in the endocardium. When the period of "ischemia" was reduced to 20 min, recovery from transmural conduction block occurred sooner and coupled beats and abnormal automaticity were detected in both epicardial and endocardial layers. Superfusion with PC during both "ischemia" and reperfusion (PC1 group), or during reperfusion only (PC2 group), significantly altered the response of the preparations to reperfusion. Following 40 min "ischemia", preparations treated with PC recovered from transmural conduction block more rapidly (PC1 group, 4 min, P less than 0.05; PC2 group, 23 min, ns), compared to control.(ABSTRACT TRUNCATED AT 250 WORDS)

We examined phosphatidylcholine (PC) effects on the isolated rat heart subjected to low- or zero-flow ischemia followed by reperfusion. Untreated hearts subjected to 30 min of low-flow ischemia recovered 15% contractility following reperfusion compared to time-control hearts. Phosphatidylcholine (0.005%) addition either 10 or 20 min before ischemia significantly enhanced recovery to approximately 61% and reduced the incidence of arrhythmias during ischemia and reperfusion. Contracture during ischemia and reperfusion was significantly reduced when PC was added 20 min before ischemia. Phosphatidylcholine was ineffective when administered at the time of reperfusion except for a moderate reduction in arrhythmia development. Phosphatidylcholine also produced a salutary effect when added 20 min prior to zero-flow ischemia. Subsarcolemmal mitochondria (SLM) and, to a much lesser degree, interfibrillar mitochondria (IFM) of untreated hearts subjected to low-flow ischemia and reperfusion exhibited depressed oxidative phosphorylation which was prevented by PC. Both mitochondrial populations exhibited a marked depression in ADP/ATP translocase activity; however, this was generally unaffected by PC. Subsarcolemmal mitochondria but not IFM of zero-flow ischemic reperfused hearts also exhibited significantly depressed oxidative phosphorylation, which was unaffected by PC. Zero-flow ischemia produced a rapid and total cessation of contractility. Both populations exhibited a substantial PC-insensitive reduction in translocase activity. Our results demonstrate, for the first time, a protection by PC on the reperfused ischemic heart. The PC-induced protection following low-flow but not zero-flow ischemia is associated with improved SLM oxidative phosphorylation suggesting dissimilar contribution of mitochondria to reperfusion-associated myocardial injury.

Initial Polytron treatment with subsequent exposure to the bacterial proteinase Nagarse has been shown to result in the isolation of two distinct populations of cardiac mitochondria, subsarcolemmal and interfibrillar mitochondria, respectively. Although these populations have been shown to possess distinct biochemical properties, few studies have been reported which document the potential differences in their response to pathological insult. We therefore examined the effect of acute hypoxia with or without reoxygenation as well as treatment with phosphate on oxidative phosphorylation on both groups of mitochondria. Freshly-isolated interfibrillar mitochondria (IFM) exhibited significantly higher respiratory values, with the exception of the ADP:O ratios, than subsarcolemmal mitochondria (SLM). With pyruvate-malate as respiratory substrate, 40 minutes hypoxia alone produced no effect on SLM whereas a stimulation in respiration was seen in IFM. A 40-minute reoxygenation period depressed the oxidative phosphorylation rate in SLM whereas it was stimulated in IFM. These treatments did not produce any effect in either population when succinate was the substrate of choice. Because of the latter observation, the possibility that increased lability of complex I of the electron transport chain accounted for the differences associated with NAD-linked substrates was studied by assessing NADH oxidation of sonicated mitochondria following the treatments. SLM exhibited enhanced permeability to exogenous NADH as well as increased sensitivity to sonication following either hypoxia or hypoxia/reoxygenation compared to IFM. Compared to hypoxia/reoxygenation, increasing concentrations of phosphate (5-15 mM) produced a marked depression in oxidative phosphorylation of SLM whereas IFM were relatively resistant. The toxic effects of phosphate were much more evident with pyruvate-malate as substrates; with succinate, oxidative phosphorylation of IFM was not depressed by phosphate whereas only a slight depression was observed with SLM. The latter population similarly exhibited reduced NADH oxidation following phosphate treatment whereas IFM were unaffected. Our studies show a differential sensitivity of two mitochondrial populations to hypoxia/reoxygenation, and, more markedly to phosphate. Since these effects were much less pronounced with succinate-linked respiration and since they were associated with defective NADH oxidation in SLM, it is suggested that the differences between the two populations may be accounted for by the increased lability of complex I of SLM due to hypoxia/reoxygenation or phosphate.

The effects of interleukin 1 (IL-1) on induction and maintenance of immune tolerance in vitro have been studied by using splenocytes from C57BL/6 male mice. B cell tolerance to the hapten trinitrophenol (TNP) was induced with TNP-human gamma globulin (HGG) and the cells were challenged with TNP-Ficoll. To determine the tolerance (or immunity), antibody concentrations in the supernatant fluids were measured by using a TNP-specific ELISA assay. Partially purified murine IL-1 abrogated tolerance induction, and when it was added at challenge phase, it also abrogated tolerance. In addition, partially purified IL-1 converted TNP-HGG from a tolerogen into an immunogen without any additional exposure to antigen. Similar results were obtained when recombinant human IL-1 alpha was used in place of partially purified natural IL-1. IL-1 is most likely acting directly on B cells rather than through the agency of T cells because purified B cells failed to become tolerant in the presence of IL-1. Studies of IL-1 production by antigen- or tolerogen-stimulated splenocytes or purified B cells showed that only antigen could elicit IL-1 production in these cells. That tolerance abrogation is unique to IL-1 is suggested by studies which show that TNF, IL-2, and INF gamma, alone, in combination with each other, or in combination with subeffective concentrations of IL-1 failed to effect tolerance abrogation.

Phosphate (Pi)-induced depression in cardiac mitochondrial function was studied using mitochondria isolated by two different procedures which purportedly yield two distinct populations. Subsarcolemmal mitochondria (SLM) exhibited an enhanced sensitivity to 20 mM Pi with respect to oxidative phosphorylation. Thus, a significant depression in oxidative phosphorylation in this population was seen following only 1-min treatment, whereas interfibrillar mitochondria (IFM) were unaffected. Both populations showed a similar response to 5-min treatment with Pi. The Pi-induced depression in respiration was partially, although significantly, reversed by a 50 microM concentration of the calcium antagonist verapamil, an observation which suggests a contribution of calcium to the Pi-induced defect in respiration. Pi also produced a potent inhibition of ADP uptake in both mitochondrial populations, which was in close agreement to Pi-induced modification of low amplitude shrinkage-swelling responses following ADP addition. Both of these parameters were unaffected by verapamil. Our results show an enhanced sensitivity of SLM to a verapamil-sensitive Pi-induced depression in oxidative phosphorylation. However, the potent, verapamil-insensitive decrease in adenine nucleotide translocase activity by Pi demonstrates that calcium is likely only partially involved in Pi-induced depression in oxidative phosphorylation and that a further partial contribution arises from a decrease in adenine nucleotide translocase activity.

1. The effect of 100 microM (20 micrograms ml-1) of D,L-carnitine was studied on the isolated heart of the rat subjected to 30 min of low flow ischaemia followed by reperfusion. 2. In untreated hearts (n = 30) ischaemia produced an almost total loss of contractility (P less than 0.05 compared with non-ischaemic time control) which was accompanied by an increase in resting tension of approximately 235% (P less than 0.05). Ventricular arrhythmias developed during ischaemia in 100% (P less than 0.05) of untreated hearts studied. Following reperfusion, untreated hearts recovered 16.3% of contractile function and demonstrated a 60% elevation in resting tension. The incidence of reperfusion-associated ventricular fibrillation was 60%. 3. Carnitine treatment produced no effect on either the contractile depression or the elevation in resting tension during ischaemia but did significantly decrease the incidence of arrythmias at the termination of ischaemia to 63.3% (n = 30, P less than 0.05). In the presence of carnitine, contractile recovery at the end of reperfusion was significantly increased to 30.2% (n = 10, P less than 0.05) and the elevation in resting tension was decreased to 30% (n = 10, P greater than 0.05). The incidence of ventricular arrhythmias during reperfusion was significantly reduced by carnitine. 4. Two populations of mitochondria, subsarcolemmal (SLM) and interfibrillar (IFM) isolated at the end of the ischaemic period exhibited an overall increase in oxidative phosphorylation rates as well as uncoupled oxygen consumption; both phenomena were more pronounced with IFM. Carnitine generally potentiated this response. A 29% and 38% inhibition in atractyloside-sensitive ADP uptake was observed in SLM and IFM, respectively, following ischaemia, which was partially prevented by carnitine. 5. After 10min of reperfusion, adenosine diphosphate (ADP) uptake in SLM was further reduced to 55% of control whereas with IFM, uptake was not different from that seen at the end of ischaemia. Mitochondria isolated from hearts after 30 min of reperfusion revealed a significantly depressed oxidative phosphorylation as well as ADP/ATP translocase activity. These defects were partially reversed in hearts perfused with carnitine. 6. Our study demonstrates that D,L-carnitine protects the rat isolated heart against injury associated with ischaemia and reperfusion through a mechanism associated with improved mitochondrial function.