A number of studies have been published on outcomes of cancer patients infected with the SARS-CoV-<em>2</em> virus causing the COVI<em>D</em>-19 infection. However, most of these reports are single-center studies with a limited number of patients. To better assess the outcomes of this new infection in this subgroup of susceptible patients, we performed a systematic review and meta-analysis to evaluate the impact of COVI<em>D</em>-19 infection on cancer patients. We searched PubMed, Web of Science, and Scopus for studies that reported the risk of infection and complications of COVI<em>D</em>-19 in cancer patients. The literature search retrieved <em>2</em><em>2</em> studies (1018 cancer patients). The analysis showed that the frequency of cancer among COVI<em>D</em>-19 confirmed patients was <em>2</em>.1% (95% CI: 1.3%, 3%) in the overall cohort. These patients had a mortality of <em>2</em>1.1% (95% CI: 14.7%, <em>2</em>7.6%), severe/critical disease rate of 45.4% (95% CI: 37.4%, 53.3%), ICU admission rate of 14.5% (95% CI: 8.5%, <em>2</em>0.4%), and mechanical ventilation rate of 11.7% (95% CI: 5.5%, 18%). The double-arm analysis showed that cancer patients had higher risk of mortality (OR = 3.<em>2</em>3, 95% CI: 1.71, 6.13), severe/critical disease (OR = 3.91, 95% CI: <em>2</em>.70, 5.67), ICU admission (OR = 3.10, 95% CI: 1.85, 5.17), and mechanical ventilation (OR = 4.86, 95% CI: 1.<em>2</em>7, 18.65), compared to non-cancer patients. Further, cancer patients had significantly lower platelet levels and a significantly higher <em>D</em>-<em>dimer</em>, C-reactive protein, and prothrombin time. In conclusion, these results indicate that cancer patients are at a higher risk of COVI<em>D</em>-19 infection-related complications. Therefore, cancer patients need diligent preventive care measures and aggressive surveillance for earlier detection of COVI<em>D</em>-19 infection.

Keywords: COVID-19; Cancer; Mechanical ventilation; Meta-analysis; Mortality.

<strong class="sub-title"> Background: </strong> As of 8 April <em>2</em>0<em>2</em>1, a total of <em>2</em>.9 million people have died with or from the coronavirus infection causing COVI<em>D</em>-19 (Corona Virus <em>D</em>isease <em>2</em>019). On <em>2</em>9 January <em>2</em>0<em>2</em>1, the European Medicines Agency (EMA) approved a COVI<em>D</em>-19 vaccine developed by Oxford University and AstraZeneca (AZ<em>D</em>1<em>2</em><em>2</em><em>2</em>, ChAdOx1 nCoV-19, COVI<em>D</em>-19 vaccine AstraZeneca, Vaxzevria, Covishield). While the vaccine prevents severe course of and death from COVI<em>D</em>-19, the observation of pulmonary, abdominal, and intracranial venous thromboembolic events has raised concerns.

Objective: To describe the clinical manifestations and the concerning management of patients with cranial venous sinus thrombosis following first exposure to the "COVID-19 vaccine AstraZeneca".

Methods: Patient files, laboratory findings, and diagnostic imaging results, and endovascular interventions of three concerning patients were evaluated in retrospect.

<strong class="sub-title"> Results: </strong> Three women with intracranial venous sinus thrombosis after their first vaccination with "COVI<em>D</em>-19 vaccine AstraZeneca" were encountered. Patient #1 was <em>2</em><em>2</em> years old and developed headaches four days after the vaccination. On day 7, she experienced a generalized epileptic seizure. Patient #<em>2</em> was 46 years old. She presented with severe headaches, hemianopia to the right, and mild aphasia 13 days after the vaccination. MRI showed a left occipital intracerebral hemorrhage. Patient #3 was 36 years old and presented 17 days after the vaccination with acute somnolence and right-hand hemiparesis. The three patients were diagnosed with extensive venous sinus thrombosis. They were managed by heparinization and endovascular recanalization of their venous sinuses. They shared similar findings: elevated levels of <em>D</em>-<em>dimers</em>, platelet factor 4 antiplatelet antibodies, corona spike protein antibodies, combined with thrombocytopenia. Under treatment with low-molecular-weight heparin, platelet counts normalized within several days.

Conclusion: Early observations insinuate that the exposure to the "COVID-19 vaccine AstraZeneca" might trigger the expression of antiplatelet antibodies, resulting in a condition with thrombocytopenia and venous thrombotic events (e.g., intracranial venous sinus thrombosis). These patients' treatment should address the thrombo-embolic manifestations, the coagulation disorder, and the underlying immunological phenomena.

Keywords: COVID-19 vaccine AstraZeneca; anticoagulation; platelet factor 4 antibodies; rheolysis; venous sinus thrombosis.

The 19 S regulatory complex (RC) of the <em>2</em>6 S proteasome is composed of at least 18 different subunits, including six ATPases that form specific pairs S4-S7, S6-S8, and S6'-S10b in vitro. One of the largest regulatory complex subunits, S<em>2</em>, was translated in reticulocyte lysate containing [(35)S]methionine and used to probe membranes containing S<em>D</em>S-polyacrylamide gel electrophoresis separated RC subunits. S<em>2</em> bound to two ATPases, S4 and S7. Association of S<em>2</em> with regulatory complex subunits was also assayed by co-translation and sedimentation. S<em>2</em> formed an immunoprecipitable heterotrimer upon co-translation with S4 and S7. The non-ATPase S5b also formed a ternary complex with S4 and S7 and the three proteins assembled into a tetramer with S<em>2</em>. Neither S<em>2</em> nor S5b formed complexes with S6'-S10b <em>dimers</em> or with S6-S8 oligomers. The use of chimeric ATPases demonstrated that S<em>2</em> binds the NH(<em>2</em>)-terminal region of S4 and the COOH-terminal two-thirds of S7. Conversely, S5b binds the COOH-terminal two-thirds of S4 and to S7's NH(<em>2</em>)-terminal region. The demonstrated association of S<em>2</em> with ATPases in the mammalian 19 S regulatory complex is consistent with and extends the recent finding that the yeast RC is composed of two subcomplexes, the lid and the base (Glickman, M. H., Rubin, <em>D</em>. M., Coux, O., Wefes, I., Pfeifer, G., Cejka, Z., Baumeister, W., Fried, V. A., and Finley, <em>D</em>. (1998) Cell 94, 615-6<em>2</em>3).

Nephrotic syndrome is associated with numerous blood coagulation abnormalities and a marked propensity to thromboembolism. The present study was undertaken to examine the status of the fibrinolytic system in this hypercoagulable state. We measured the antigen concentrations or activities of plasminogen, tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor (PAI), alpha <em>2</em>-antiplasmin, alpha 1-antitrypsin, and alpha <em>2</em>-macroglobulin as well as total antiplasmin activity and <em>D</em>-<em>dimer</em> concentration in the plasma of 39 patients with nephrotic syndrome and 3<em>2</em> normal controls subjects. In addition, antigen concentrations of plasminogen, alpha <em>2</em>-antiplasmin, alpha 1-antitrypsin, and alpha <em>2</em>-macroglobulin were measured in the urine of the study populations. The nephrotic group showed marked elevations of plasma t-PA, plasminogen, alpha <em>2</em>-macroglobulin, and <em>D</em>-<em>dimer</em> and a significant reduction of plasma alpha <em>2</em>-antiplasmin and alpha 1-antitrypsin as compared with the normal control group. Plasma alpha <em>2</em>-macroglobulin was directly related to <em>2</em>4-hour urinary protein excretion and inversely related to serum albumin concentration. None of the proteins measured were detectable in the urine of normal controls. However, substantial amounts of plasminogen, alpha <em>2</em>-antiplasmin, and alpha 1-antitrypsin and small amounts of alpha <em>2</em>-macroglobulin were recovered in the urine of patients with nephrotic syndrome. <em>D</em>espite the lack of clinically demonstrable thrombosis, plasma <em>D</em>-<em>dimer</em> was markedly elevated in the nephrotic group, suggesting concurrent activation of coagulation and fibrinolytic pathways. In addition, the study revealed multiple abnormalities of the plasma fibrinolytic proteins and documented their urinary excretion in patients with nephrotic syndrome.(ABSTRACT TRUNCATE<em>D</em> AT <em>2</em>50 WOR<em>D</em>S)

OBJECTIVE

To determine the degree of regional and systemic coagulation activation soon after isolated severe head injury.

METHODS

Prospective, controlled clinical study.

METHODS

The emergency room and intensive care unit (ICU) of a trauma center in a university hospital serving a population of 5 million people.

METHODS

Twenty-four trauma victims: <em>2</em>0 patients with isolated severe head injury (head trauma group, Glasgow Coma Score of < or =8) and four patients with isolated bone fracture (control group).

METHODS

Insertion of central venous, i.e. superior caval vein, jugular bulb, and arterial catheters for blood sampling.

RESULTS

Central venous (i.e., superior caval vein) global coagulation variables (i.e., prothrombin time, activated partial thromboplastin time, fibrinogen concentration, antithrombin III activity, and platelet count) and central venous and jugular bulb activation coagulation and fibrinolysis variables (i.e., prothrombin fragment F1+<em>2</em>, thrombin-antithrombin III complex, soluble fibrin, and <em>D</em>-<em>dimer</em> concentration) were measured soon after trauma (<6 hrs) and 3 hrs later. When compared with the control group patients, upon ICU admission, fibrinogen concentration (p < .005) and platelet count (p < .0<em>2</em>5) were significantly decreased in the head trauma group. Thrombin-antithrombin III complex (p < .0<em>2</em>5), prothrombin fragment F1+<em>2</em> (p < .0<em>2</em>5), and <em>D</em>-<em>dimer</em> (p < .005) concentrations measured at the time of ICU admission were significantly higher in the head trauma group than in the control group. Only in the head trauma group were soluble fibrin concentrations increased above the normal range; thrombin-antithrombin III complex and the prothrombin fragment F1+<em>2</em> were found to be significantly higher in cerebrovenous blood than in central venous blood (p < .0<em>2</em>5). There was no cerebrocentral venous difference in the concentrations of soluble fibrin or <em>D</em>-<em>dimer</em> in either group.

CONCLUSIONS

Within 6 hrs after severe isolated head trauma, systemic procoagulant overflow from the traumatized cerebral microvasculature proceeds to the thrombin level and is then inhibited by antithrombin III. Regional and systemic hypercoagulability and increased D-dimer concentrations appear to be common among head trauma patients. Increased procoagulant and consecutive fibrinolytic turnover may, therefore, spark disseminated intravascular coagulation in this patient group.

OBJECTIVE

The aim of this study was to evaluate the relation between erectile dysfunction and endothelial functions, coagulation activation, peripheral and autonomic neuropathy in men with Type II (non-insulin-dependent) diabetes mellitus.

METHODS

We studied 30 Type II diabetic patients with symptomatic erectile dysfunction and 30 potent diabetic patients matched for age and disease. Endothelial functions were assessed with the L-arginine test, plasma thrombomodulin and cell adhesion molecules circulating concentrations. Haemostasis was evaluated with markers of thrombin activation and fibrinolysis. Quantitative sensory testing (vibratory, warming, and heat-pain thresholds), cardiovascular reflex tests and <em>2</em>4-h blood pressure monitoring were used to assess peripheral or autonomic neuropathy.

RESULTS

Mean erectile score and HbA1c were 10.5+/-5.8 and 8.3+/-1.6% in patients with erectile dysfunction, and <em>2</em>4.0+/-0.7 and 6.8+/-1.4% in those without erectile dysfunction, respectively (p < 0.001); there was a significant relation between HbA1c and erectile function score in patients with erectile dysfunction (r = -0.45, p = 0.0<em>2</em>). The decrease in blood pressure and platelet aggregation in response to L-arginine was lower (p < 0.05-0.0<em>2</em>) in patients with erectile dysfunction, whereas soluble thrombomodulin, P-selectin and intercellular cell ahhesion molecule-1 concentrations were higher (p < 0.05-0.0<em>2</em>). Indices of coagulation activation (F1 + <em>2</em> and <em>D</em>-<em>dimers</em>) and reduced fibrinolysis (PAI-1) were also found to be higher in erectile dysfunction patients. Heat-pain and warm perception thresholds. as well as cardiovascular reflex tests, were most commonly abnormal in patients with erectile dysfunction (p < 0.05). In multivariate analysis, HbA1c, MBP response to L-arginine, P-selectin, indices of coagulation, and quantitative sensory testing were independent predictors of erectile function score.

CONCLUSIONS

Erectile dysfunction in diabetic men correlates with endothelial dysfunction. A reduced nitric oxide activity might provide a unifying explanation.

AIMS AND BACKGROUNDS: Plasma concentrations of several proteases of the coagulation system have been shown to predict prognosis in malignancy. The study was aimed to investigate the prognostic value of plasma D-dimer concentrations and some other coagulation factors in lung cancer.

METHODS

Between 2004 and 2008, 100 newly diagnosed lung cancer patients and 25 healthy individuals serving as the control group were evaluated. The patients had no history of coagulation system disorders or anticoagulant therapy. Plasma D-dimer concentrations, prothrombin time, activated partial thromboplastin time, international normalized ratio and blood counts of the patients were obtained. Patient age, lung cancer stage, tumor histology, therapy modalities (surgery, chemotherapy and radiotherapy), therapy outcomes and survival durations of the patients were determined.

RESULTS

The median age of the patients (86 males/14 females) was 67 years, and 15% had stage 2, 26% had stage 3A, 24% had stage 3B, and 35% had stage 4 disease. Histologic subtypes were non-small cell carcinoma (87%) and small cell carcinoma (13%). The median D-dimer level of the patients was 1250 ng/dl, which was significantly higher than that of the control group. Survival duration was significantly higher in patients with low D-dimer levels (P <0.05). D-dimer plasma levels predicted survival independently of the clinical stage of disease, histologic tumor type and performance status of the patient (HR = 5.1; 95% confidence interval, 1.015-1.19, P = 0.013). Plasma D-dimer level was significantly higher in metastatic disease (P <0.01).

CONCLUSIONS

The results suggest that D-dimer plasma levels might be useful to predict the clinical outcome and survival of patients with lung cancer.

DNA from the telomeres at the en<em>d</em>s of eukaryotic chromosomes contains a stretch of simple tan<em>d</em>emly repeate<em>d</em> sequences in which clusters of G resi<em>d</em>ues alternate with clusters of T/A sequences along one DNA stran<em>d</em>. Mo<em>d</em>el telomeric G-clusters form four-stran<em>d</em>e<em>d</em> structures in Na+ or K+, stabilize<em>d</em> by Hoogsteen pairing between G bases. DNA containing a single copy of the G-cluster can self-associate to form tetramers, with a parallel-stran<em>d</em>e<em>d</em>, right-han<em>d</em>e<em>d</em> helical structure. Two copies of the 3'-terminal G stran<em>d</em> form a fol<em>d</em>e<em>d</em>-back hairpin that <em>d</em>imerizes to create an antiparallel qua<em>d</em>ruplex structure. We show here that the tetrameric structure is strongly influence<em>d</em> by the T resi<em>d</em>ue flanking either si<em>d</em>e of the G-cluster. The parallel tetraplex forme<em>d</em> by single copies of the sequences <em>d</em>TnG4 is most stable for n = 1 an<em>d</em> least stable for n = 8, the longest tract we have stu<em>d</em>ie<em>d</em>. At least two thymine resi<em>d</em>ues are require<em>d</em> to allow formation of antiparallel fol<em>d</em>e<em>d</em>-back hairpin <em>dimers</em> from two-copy oligomers of sequence <em>d</em>(TnG4)<em>2</em> in Na+; a<em>d</em><em>d</em>itional T's <em>d</em>estabilize this structure. In K+, the pre<em>d</em>ominant structure forme<em>d</em> is the four-stran<em>d</em>e<em>d</em> parallel tetramer in all cases. Kinetic analysis in<em>d</em>icates that the qua<em>d</em>ruplex structure forme<em>d</em> by Oxytricha telomeric DNA overhangs in the presence of Na+ arises by <em>d</em>imerization of two Hoogsteen base-paire<em>d</em> hairpins, with a relatively low energy barrier.

The fin<em>d</em>ing that exchange of tubulin subunits between tubulin <em>dimers</em> (alpha-beta + alpha'beta' <->> alpha'beta + alphabeta') <em>d</em>oes not occur in the absence of protein cofactors an<em>d</em> GTP hy<em>d</em>rolysis conflicts with the assumption that pure tubulin <em>dimer</em> an<em>d</em> monomer are in rapi<em>d</em> equilibrium. This assumption un<em>d</em>erlies the many physical chemical measurements of the K(<em>d</em>) for <em>dimer</em> <em>d</em>issociation. To resolve this <em>d</em>iscrepancy we use<em>d</em> surface plasmon resonance to <em>d</em>etermine the rate constant for <em>dimer</em> <em>d</em>issociation. The half-time for <em>d</em>issociation was approximately 9.6 h with tubulin-GTP, <em>2</em>.4 h with tubulin-GDP, an<em>d</em> 1.3 h in the absence of nucleoti<em>d</em>e. A K<em>d</em> equal to 10(-11) M was calculate<em>d</em> from the measure<em>d</em> rate for <em>d</em>issociation an<em>d</em> an estimate<em>d</em> rate for association. <em>Dimer</em> <em>d</em>issociation was foun<em>d</em> to be reversible, an<em>d</em> <em>dimer</em> formation <em>d</em>oes not require GTP hy<em>d</em>rolysis or fol<em>d</em>ing information from protein cofactors, because 0.<em>2</em> microM tubulin-GDP incubate<em>d</em> for <em>2</em>0 h was elute<em>d</em> as <em>dimer</em> when analyze<em>d</em> by size exclusion chromatography. Because <em>2</em>0 h correspon<em>d</em>s to eight half-times for <em>d</em>issociation, only monomer woul<em>d</em> be present if <em>d</em>issociation were an irreversible reaction an<em>d</em> if <em>dimer</em> formation require<em>d</em> GTP or protein cofactors. A<em>d</em><em>d</em>itional evi<em>d</em>ence for a 10(-11) M K(<em>d</em>) was obtaine<em>d</em> from gel exclusion chromatography stu<em>d</em>ies of 0.0<em>2</em>-<em>2</em> nM tubulin-GDP. The slow <em>d</em>issociation of the tubulin <em>dimer</em> suggests that protein tubulin cofactors function to catalyze <em>dimer</em> <em>d</em>issociation, rather than <em>dimer</em> assembly. Assuming N-site-GTP <em>d</em>issociation is from monomer, our results agree with the 16-h half-time for N-site GTP in vitro an<em>d</em> 33 h half-life for tubulin N-site-GTP in CHO cells.

Interactions between the C-terminal interface residues (96-99) of the mature HIV-1 protease were shown to be essential for <em>dimer</em>ization, whereas the N-terminal residues () and Arg(87) contribute to <em>dimer</em> stability (Ishima, R., Ghirlando, R., Tozser, J., Gronenborn, A. M., Torchia, <em>D</em>. A., and Louis, J. M. (<em>2</em>001) J. Biol. Chem. <em>2</em>76, 49110-49116). Here we show that the intramonomer interaction between the side chains of Asp(<em>2</em>9) and Arg(87) influences <em>dimer</em>ization significantly more than the intermonomer interaction between Asp(<em>2</em>9) and Arg(8'). Several mutants, including T<em>2</em>6A, destablize the <em>dimer</em>, exhibit a monomer fold, and are prone to aggregation. To alleviate this undesirable property, we designed proteins in which the N- and C-terminal regions can be linked intramolecularly by disulfide bonds. In particular, cysteine residues were introduced at positions <em>2</em> and 97 or 98. A procedure for the efficient preparation of intrachain-linked polypeptides is presented, and it is demonstrated that the Q<em>2</em>C/L97C variant exhibits a native-like single subunit fold. It is anticipated that monomeric proteases of this kind will aid in the discovery of novel inhibitors aimed at binding to the monomer at the <em>dimer</em>ization interface. This extends the target area of current inhibitors, all of which bind across the active site formed by both subunits in the active <em>dimer</em>.

BACKGROUND

Intravascular fibrin deposition and platelet sequestration occur with porcine xenograft rejection by baboons. Disseminated intravascular coagulopathy may arise either as a direct consequence of the failure to fully deplete xenoreactive natural antibodies and block complement, or because of putative cross-species molecular incompatibilities in this discordant species combination.

METHODS

Three baboons were conditioned with retrovirally transduced autologous bone marrow to induce tolerance to swine antigens. Xenoreactive natural antibodies and complement were depleted by plasmapheresis and the use of Gal alpha1-3Gal column adsorptions; baboons were then splenectomized and underwent renal xenografting from inbred, miniature pigs. Soluble complement receptor type-1 with protocol immunosuppression (mycophenolate mofetil, 15-deoxyspergualin, steroids, and cyclosporine) was administered.

RESULTS

A bleeding diathesis was clinically evident from days 5 to 1<em>2</em> after transplantation in two baboons. Low levels of circulating C3a, C3d, and iC3b were measured despite the absence of functional circulating complement components. Profound thrombocytopenia with abnormalities in keeping with disseminated intravascular coagulopathy were observed. Prolongation of prothrombin and partial thromboplastin times was accompanied by evidence for tissue factor-mediated coagulation pathways, high levels of thrombin generation (prothrombin fragment F(1+<em>2</em>) production and thrombin-antithrombin complex formation), fibrinogen depletion, and production of high levels of the fibrin degradation product <em>D</em>-<em>dimer</em>. Importantly, these disturbances resolved rapidly after the excision of the rejected xenografts in two surviving animals. Histopathological examination of the rejected xenografts confirmed vascular injury, fibrin deposition, platelet deposition, and localized complement activation.

CONCLUSIONS

Systemic coagulation disturbances are associated with delayed xenograft rejection.

Mitochondria from plants, yeast, and animals each contain at least one peroxiredoxin (Prx) that is involved in peroxide detoxification and redox signalling. The supramolecular dynamics of atypical type II Prx targeted to the mitochondrion was addressed in pea. Microcalorimetric (ITC) titrations identified an extremely high-affinity binding between the mitochondrial PsPrxIIF and Trx-o with a K(<em>D</em>) of 1<em>2</em>6+/-14 pM. Binding was driven by a favourable enthalpy change (<em>D</em>eltaH= -60.6 kcal mol(-1)) which was counterbalanced by unfavourable entropy changes (T<em>D</em>eltaS= -47.1 kcal mol(-1)). This is consistent with the occurrence of large conformational changes during binding which was abolished upon site-directed mutaganesis of the catalytic C59S and C84S. The redox-dependent interaction was confirmed by gel filtration of mitochondrial extracts and co-immunoprecipitation from extracts. The heterocomplex of PsPrxIIF and Trx-o reduced peroxide substrates more efficiently than free PsPrxIIF suggesting that Trx-o serves as an efficient and specific electron donor to PsPrxIIF in vivo. Other Trx-s tested by ITC analysis failed to interact with PsPrxIIF indicating a specific recognition of PsPrxIIF by Trx-o. PsPrxIIF exists primarily as a <em>dimer</em> or a hexamer depending on the redox state. In addition to the well-characterized oligomerization of classical <em>2</em>-Cys Prx the results also show that atypical Prx undergo large structural reorganization with implications for protein-protein interaction and function.

The Fur apoprotein has been purified and reconstituted with Co<em>2</em>+ and Mn<em>2</em>+ ions. These samples have been analyzed by UV-visible, EPR, and 1H NMR spectroscopies, by XAS, and by magnetization measurements. The apo-Fur protein is able to bind one metal dication (Co<em>2</em>+ or Mn<em>2</em>+) per monomer. A saturation magnetization study confirms the presence of a high-spin metal dication [Mn(II) S = 5/<em>2</em> and Co(II) S = 3/<em>2</em>]. The two metal ions per Fur <em>dimer</em> are not in magnetic interaction (|J| < 0.1 cm-1 ). The UV-visible spectrum of the cobalt-substituted form (Co-Fur) presents two main bands at 660 nm and 540(br) nm with epsilon540 nm = 65 M-1 cm-1. The EPR spectrum gives the following g values: gx = 5.0(5), gy = 4.0(<em>2</em>), and gz = <em>2</em>. 3(1), which are in accordance with a nearly axial (E/<em>D</em> < 0.11) site. The value of 55 cm-1 for the splitting (<em>Delta</em>) between the ground and the first excited state has been derived from an EPR saturation study and is in agreement with magnetization data. The EXAFS data of Co-Fur indicate a metal environment comprising five nitrogen/oxygen atoms at <em>2</em>.11 A, the absence of sulfur, and the presence of histidines as ligands. 1H NMR of Co-Fur in H<em>2</em>O and <em>D</em><em>2</em>O shows at least two exchangeable signals coming from histidine NH protons and shows the signature of carboxylate group(s). The combined spectroscopic data allow us to propose that the main metal site of Fur in Co-Fur contains at least two histidines, at least one aspartate or glutamate, and no cysteine as ligands and is in an axially distorted octahedral environment.

Recombinant human brain calbindin <em>D</em>(<em>2</em>8K) (rHCaBP), human Cu,Zn-superoxide dismutase (HCuZnSO<em>D</em>), rabbit muscle glyceraldehyde-3-phosphate dehydrogenase (GAP<em>D</em>H), and bovine serum albumin (BSA) were found to be S-glutathiolated in decomposed S-nitrosoglutathione (GSNO) solutions. Tryptic or Glu-C digestion and MAL<em>D</em>I-TOF MS analyses of the digests are consistent with S-thiolation of Cys111 and Cys187 of HCuZnSO<em>D</em> and rHCaBP, respectively, upon exposure to decomposed GSNO. GAP<em>D</em>H activity analysis reveals that S-glutathiolation most likely occurs on the active site Cys149, and the single free Cys34 is assumed to be the site of S-glutathiolation in BSA. The yields of S-glutathiolation of rHCaBP, GAP<em>D</em>H, and BSA were much higher than those of HCuZnSO<em>D</em>. The latter is limited by the accessibility of Cys111 to the glutathiolating reagent in the HCuZnSO<em>D</em> <em>dimer</em>. Unlike decomposed GSNO, fresh GSNO, reduced glutathione (GSH), and oxidized glutathione (GSSG) are not efficient S-glutathiolating agents for the proteins examined here. On the basis of analysis by mass spectrometry and UV-visible absorption, GSNO decomposition in the dark at room temperature yields glutathione disulfide S-oxide [GS(O)SG], glutathione disulfide S-dioxide (GSO(<em>2</em>)SG), and GSSG as products. GS(O)SG is the efficient protein S-glutathiolating agent in GSNO solutions, not GSNO, which does not carry out efficient S-glutathiolation of rHCaBP, HCuZnSO<em>D</em>, or GAP<em>D</em>H in vitro. A hydrolysis pathway yielding GSOH and nitroxyl (HNO/NO(-)) as intermediates is proposed for GSNO decomposition in the dark. This is based on inhibition of GSNO breakdown by dimedone, a reagent specific for sulfenic acids, and on nitroxyl scavenging by metmyoglobin. The results presented here are contrary to numerous reports of protein S-thiolation by low-molecular weight S-nitrosothiols.

NHE3 activity is regulated by phosphorylation/dephosphorylation processes and membrane recycling in intact cells. However, the Na(+)/H(+) exchanger (NHE) can also be regulated by G proteins independent of cytoplasmic second messengers, but the G protein subunits involved in this regulation are not known. Therefore, we studied G protein subunit regulation of NHE3 activity in renal brush-border membrane vesicles (BBMV) in a system devoid of cytoplasmic components and second messengers. Basal NHE3 activity was not regulated by G(s)alpha or G(i)alpha, because antibodies to these G proteins by themselves were without effect. The inhibitory effect of <em>D</em>(1)-like agonists on NHE3 activity was mediated, in part, by G(s)alpha, because it was partially reversed by anti-G(s)alpha antibodies. Moreover, the amount of G(s)alpha that coimmunoprecipitated with NHE3 was increased by fenoldopam in both brush-border membranes and renal proximal tubule cells. Furthermore, guanosine 5'-O-(3-thiotriphosphate) but not guanosine 5'-O-(<em>2</em>-thiodiphosphate), the inactive analog of G<em>D</em>P, increased the amount of G(s)alpha that coimmunoprecipitated with NHE3. The alpha(<em>2</em>)-adrenergic agonist, UK-14304 or pertussis toxin (PTX) alone had no effect on NHE3 activity, but UK-14304 and PTX treatment attenuated the <em>D</em>(1)-like receptor-mediated NHE3 inhibition. The ability of UK-14304 to attenuate the <em>D</em>(1)-like agonist effect was not due to G(i)alpha, because the attenuation was not blocked by anti-G(i)alpha antibodies or by PTX. Anti-Gbeta(common) antibodies, by themselves, slightly inhibited NHE3 activity but had little effect on <em>D</em>(1)-like receptor-mediated NHE3 inhibition. However, anti-Gbeta(common) antibodies reversed the effects of UK-14304 and PTX on <em>D</em>(1)-like agonist-mediated NHE3 inhibition. These studies provide concrete evidence of a direct regulatory role for G(s)alpha, independent of second messengers, in the <em>D</em>(1)-like-mediated inhibition of NHE3 activity in rat renal BBMV. In addition, beta/gamma <em>dimers</em> of heterotrimeric G proteins appear to have a stimulatory effect on NHE3 activity in BBMV.

In this study we addressed the questions as to whether repair is confined to the nuclear matrix compartment, analogous to replication and transcription and how repair events are distributed in <em>D</em>NA loops associated with the nuclear matrix. Pulse labelling of ultraviolet (<em>2</em>54 nm) irradiated confluent human fibroblasts revealed that repair was preferentially located in nuclear matrix associated <em>D</em>NA in cells exposed to 5 J/m<em>2</em>. However, in cells exposed to 30 J/m<em>2</em> repair approached a random distribution. The non-random distribution of repair label at 5 J/m<em>2</em> was most pronounced directly after irradiation and gradually changed to a more random distribution within two hours after treatment. The results of pulse-chase experiments exclude the possibility of transient binding of repair sites to the matrix and favour the model of preferential repair of <em>D</em>NA sequences permanently associated with the nuclear matrix. Pronounced differences in distribution pattern of repair events in <em>D</em>NA loops were found among normal and UV-sensitive cell lines exposed to 5 J/m<em>2</em>. Repair in nuclear matrix associated <em>D</em>NA was 1.7 fold more efficient than in loop <em>D</em>NA in normal and xeroderma pigmentosum group <em>D</em> cells and over 3 fold in xeroderma pigmentosum group C cells. In Cockayne's syndrome fibroblasts repair in nuclear matrix <em>D</em>NA was found to be <em>2</em> fold less efficient than in loop <em>D</em>NA. This heterogeneity in distribution of repair correlates well with preferential removal of pyrimidine <em>dimers</em> from transcriptionally active <em>D</em>NA in normal and xeroderma pigmentosum group C cells and its absence in Cockayne's syndrome cells as recently reported by Mayne et al., 1988. The results suggest that Cockayne's syndrome cells have a defect in excision of UV-damage from transcriptionally active genes located proximal to the nuclear matrix. Xeroderma pigmentosum group C cells may possess a defect in <em>D</em>NA repair associated with chromatin regions outside transcriptionally active <em>D</em>NA.

The urease accessory protein enco<em>d</em>e<em>d</em> by ureE from Klebsiella aerogenes is propose<em>d</em> to <em>d</em>eliver Ni(II) to the urease apoprotein <em>d</em>uring enzyme activation. Native UreE possesses a histi<em>d</em>ine-rich region at its carboxyl terminus that bin<em>d</em>s several equivalents of Ni(<em>2</em>+); however, a truncate<em>d</em> form of this protein (H144*UreE) bin<em>d</em>s only <em>2</em> Ni(<em>2</em>+) per <em>dimer</em> an<em>d</em> is functionally active (Brayman, T. G., an<em>d</em> Hausinger, R. P. (1996) J. Bacteriol. 178, 5410-5416). The urease activation kinetics were stu<em>d</em>ie<em>d</em> in vivo by monitoring the <em>d</em>evelopment of urease activity upon a<em>d</em><em>d</em>ing Ni(<em>2</em>+) to spectinomycin-treate<em>d</em> Escherichia coli cells that expresse<em>d</em> the complete K. aerogenes urease gene cluster with altere<em>d</em> forms of ureE. Site-specific alterations of H144*UreE <em>d</em>ecrease the rate of in vivo urease activation, with the most <em>d</em>ramatic changes observe<em>d</em> for the H96A, H110A, D111A, an<em>d</em> H11<em>2</em>A substitutions. Notably, urease activity in cells pro<em>d</em>ucing H96A/H144*UreE was lower than cells containing a ureE <em>d</em>eletion. Prior stu<em>d</em>ies ha<em>d</em> shown that H110A an<em>d</em> H11<em>2</em>A variants each boun<em>d</em> a single Ni(<em>2</em>+) per <em>dimer</em> with elevate<em>d</em> K(<em>d</em>) values compare<em>d</em> with control H144*UreE, whereas the H96A an<em>d</em> D111A variants boun<em>d</em> <em>2</em> Ni(<em>2</em>+) per <em>dimer</em> with unperturbe<em>d</em> K(<em>d</em>) values (Colpas, G. J., Brayman, T. G., Ming, L.-J., an<em>d</em> Hausinger, R. P. (1999) Biochemistry 38, 4078-4088). To un<em>d</em>erstan<em>d</em> why cells containing the latter two proteins showe<em>d</em> re<em>d</em>uce<em>d</em> rates of urease activation, we characterize<em>d</em> their metal bin<em>d</em>ing/<em>d</em>issociation kinetics an<em>d</em> compare<em>d</em> the results to those obtaine<em>d</em> for H144*UreE. The truncate<em>d</em> protein was shown to sequentially bin<em>d</em> two Ni(<em>2</em>+) with k(1) approximately 18 an<em>d</em> k(<em>2</em>) approximately 100 M(-1) s(-1), an<em>d</em> with <em>d</em>issociation rates k(-1) approximately 3 x 10(-3) an<em>d</em> k(-<em>2</em>) approximately 10(-4) s(-1). Similar apparent rates of bin<em>d</em>ing an<em>d</em> <em>d</em>issociation were note<em>d</em> for the two mutant proteins, suggesting that altere<em>d</em> H144*UreE interactions with Ni(<em>2</em>+) <em>d</em>o not account for the changes in cellular urease activation. These conclusions are further supporte<em>d</em> by in vitro experiments <em>d</em>emonstrating that a<em>d</em><em>d</em>ition of H144*UreE to urease apoprotein activation mixtures inhibite<em>d</em> the rate an<em>d</em> extent of urease formation. Our results highlight the importance of other urease accessory proteins in assisting UreE-<em>d</em>epen<em>d</em>ent urease maturation.

Cholesterol is most often found distributed nonrandomly in the plane of the bilayer, giving rise to cholesterol-rich and -poor domains. Many of these domains are thought to be crucial for the maintenance of membrane structure and function. However, such well-characterized domains generally occur in the membranes that contain relatively large amounts of cholesterol. Cholesterol organization in membranes containing very low amounts of cholesterol has not been investigated extensively. Recent evidence from differential-scanning calorimetric studies suggest that cholesterol may not form uniform monodisperse solutions, as assumed earlier, in the membranes even at very low concentrations. Fluorescent cholesterol analogues, when chosen carefully, offer a powerful approach for studying the distribution and organization of cholesterol in membranes at low concentrations. In this paper, we have studied the organization of cholesterol in membranes at very low concentrations (up to 5 mol % of the total lipid) using a fluorescent cholesterol analogue (NB<em>D</em>-cholesterol) which is labeled with the 7-nitrobenz-<em>2</em>-oxa-1,3-diazol-4-yl (NB<em>D</em>) group at the flexible acyl chain, without any alteration in the structural features necessary for proper membrane incorporation. Our results show that NB<em>D</em>-cholesterol exhibits local organization even at very low concentrations. This is consistent with the recently suggested model of cholesterol organization in membranes at low concentrations, involving the formation of transbilayer, tail-to-tail <em>dimers</em> [Harris, J.S., Epps, <em>D</em>. E., <em>D</em>avio, S. R., & Kezdy, F.J. (1995) Biochemistry 34, 3851-3857]. The implications of such local cholesterol organization in membranes that have very low cholesterol content in vivo, such as the endoplasmic reticulum and the inner mitochondrial membrane, open up interesting possibilities.

Human erythroid spectrin <em>dimer</em> assembly is initiated by the association of a specific region near the N-terminal of beta-spectrin with a complementary region near the C-terminal of alpha-spectrin (Speicher, <em>D</em>. W., Weglarz, L., and <em>D</em>eSilva, T. M. (199<em>2</em>) J. Biol. Chem. <em>2</em>67, 14775-1478<em>2</em>). Both spectrin subunits consist primarily of tandem, 106-residue long, homologous, triple-helical motifs. In this study, the minimal region of beta-spectrin required for association with alpha-spectrin was determined using recombinant peptides. The start site (phasing) for construction of <em>dimer</em>ization competent beta-spectrin peptides was particularly critical. The beginning of the first homologous motif for both beta-spectrin and the related <em>dimer</em>ization site of alpha-actinin is approximately 8 residues earlier than most spectrin motifs. A four-motif beta-spectrin peptide (beta1-4+) with this earlier starting point bound to full-length alpha-spectrin with a Kd of about 10 nM, while deletion of these first 8 residues reduced binding nearly 10-fold. N- and C-terminal truncations of one or more motifs from beta1-4+ showed that the first motif was essential for <em>dimer</em>ization since its deletion abolished binding, but beta1+ alone could not associate with alpha-monomers. The first two motifs (beta1 <em>2</em>+) represented the minimum lateral <em>dimer</em> assembly site with a Kd of about <em>2</em>30 nM for interaction with full-length alpha-spectrin or an alpha-spectrin nucleation site recombinant peptide, alpha18-<em>2</em>1. Each additional motif increased the <em>dimer</em>ization affinity by approximately 5-fold. In addition to this strong inter-subunit <em>dimer</em> association, interactions between the helices of a single triple-helical motif are frequently strong enough to maintain a noncovalent complex after internal protease cleavage similar to the interactions thought to be involved in tetramer formation. Analysis of hydrodynamic radii of recombinant peptides containing differing numbers of motifs showed that a single motif had a Stokes radius of <em>2</em>.35 nM, while each additional motif added only 0.85 nM to the Stokes radius. This is the first direct demonstration that spectrin's flexibility arises from regions between each triple helical motif rather than from within the segment itself and suggests that current models of inter-motif connections may need to be revised.

A <em>d</em>imeric neomycin-neomycin conjugate 3 with a flexible linker, <em>2</em>,<em>2</em>'-(ethylene<em>d</em>ioxy)bis(ethylamine), has been synthesize<em>d</em> an<em>d</em> characterize<em>d</em>. <em>Dimer</em> 3 can selectively bin<em>d</em> to AT-rich DNA <em>d</em>uplexes with high affinity. Biophysical stu<em>d</em>ies have been performe<em>d</em> between 3 an<em>d</em> <em>d</em>ifferent nucleic aci<em>d</em>s with varying base composition an<em>d</em> conformation by using ITC (isothermal calorimetry), CD (circular <em>d</em>ichroism), FID (fluorescent intercalator <em>d</em>isplacement), an<em>d</em> UV (ultraviolet) thermal <em>d</em>enaturation experiments. A few conclusions can be <em>d</em>rawn from this stu<em>d</em>y: (1) FID assay with 3 an<em>d</em> polynucleoti<em>d</em>es <em>d</em>emonstrates the preference of 3 towar<em>d</em> AT-rich sequences over GC-rich sequences. (<em>2</em>) FID assay an<em>d</em> UV thermal <em>d</em>enaturation experiments show that 3 has a higher affinity for the poly(<em>d</em>A)·poly(<em>d</em>T) DNA <em>d</em>uplex than for the poly(<em>d</em>A)·<em>2</em>poly(<em>d</em>T) DNA triplex. Contrary to neomycin, 3 <em>d</em>estabilizes poly(<em>d</em>A)·<em>2</em>poly(<em>d</em>T) triplex but stabilizes poly(<em>d</em>A)·poly(<em>d</em>T) <em>d</em>uplex, suggesting the major groove as the bin<em>d</em>ing site. (3) UV thermal <em>d</em>enaturation stu<em>d</em>ies an<em>d</em> ITC experiments show that 3 stabilizes continuous AT-tract DNA better than DNA <em>d</em>uplexes with alternating AT bases. (4) CD an<em>d</em> FID titration stu<em>d</em>ies show a DNA bin<em>d</em>ing site size of 10-1<em>2</em> base pairs/<em>d</em>rug, <em>d</em>epen<em>d</em>ing upon the structure/sequence of the <em>d</em>uplex for AT-rich DNA <em>d</em>uplexes. (5) FID an<em>d</em> ITC titration between 3 an<em>d</em> an intramolecular DNA <em>d</em>uplex [<em>d</em>(5'-A(1<em>2</em>)-x-T(1<em>2</em>)-3'), x = hexaethylene glycol linker] results in a bin<em>d</em>ing stoichiometry of 1:1 with a bin<em>d</em>ing constant ∼10(8) M(-1) at 100 mM KCl. (6) FID assay using 3 an<em>d</em> 51<em>2</em> hairpin DNA sequences that vary in their AT base content an<em>d</em> placement also show a higher bin<em>d</em>ing selectivity of 3 towar<em>d</em> continuous AT-rich than towar<em>d</em> DNA <em>d</em>uplexes with alternate AT base pairs. (7) Salt-<em>d</em>epen<em>d</em>ent stu<em>d</em>ies in<em>d</em>icate the formation of three ion pairs <em>d</em>uring bin<em>d</em>ing of the DNA <em>d</em>uplex <em>d</em>[5'-A(1<em>2</em>)-x-T(1<em>2</em>)-3'] an<em>d</em> 3. (8) ITC-<em>d</em>erive<em>d</em> bin<em>d</em>ing constants between 3 an<em>d</em> DNA <em>d</em>uplexes have the following or<em>d</em>er: AT continuous, <em>d</em>[5'-G(3)A(5)T(5)C(3)-3']>> AT alternate, <em>d</em>[5'-G(3)(AT)(5)C(3)-3']>> GC-rich <em>d</em>[5'-A(3)G(5)C(5)T(3)-3']. (9) 3 bin<em>d</em>s to the AT-tract-containing DNA <em>d</em>uplex (B* DNA, <em>d</em>[5'-G(3)A(5)T(5)C(3)-3']) with 1 or<em>d</em>er of magnitu<em>d</em>e higher affinity than to a DNA <em>d</em>uplex with alternating AT base pairs (B DNA, <em>d</em>[5'-G(3)(AT)(5)C(3)-3']) an<em>d</em> with almost 3 or<em>d</em>ers of magnitu<em>d</em>e higher affinity than a GC-rich DNA (A-form, <em>d</em>[5'-A(3)G(5)C(5)T(3)-3']).

Macromolecular assemblies play critical roles in regulating cellular functions. The cysteine synthase complex (CSC), which is forme<em>d</em> by association of serine O-acetyltransferase (SAT) an<em>d</em> O-acetylserine sulfhy<em>d</em>rylase (OASS), acts as a sensor an<em>d</em> mo<em>d</em>ulator of thiol metabolism by respon<em>d</em>ing to changes in nutrient con<em>d</em>itions. Here we examine the oligomerization an<em>d</em> energetics of formation of the soybean CSC. Biophysical examination of the CSC by size exclusion chromatography an<em>d</em> se<em>d</em>imentation ultracentrifugation in<em>d</em>icates that this assembly (complex M(r) approximately 330,000) consists of a single SAT trimer (trimer M(r) approximately 110,000) an<em>d</em> three OASS <em>dimers</em> (<em>dimer</em> M(r) approximately 70,000). Analysis of the SAT-OASS interaction by isothermal titration calorimetry reveals negative cooperativity with three <em>d</em>istinct bin<em>d</em>ing events <em>d</em>uring CSC formation with K(<em>d</em>) values of 0.3, 7.5, an<em>d</em> 78 nm. The three bin<em>d</em>ing events are also observe<em>d</em> using surface plasmon resonance with comparable affinities. The stability of the CSC <em>d</em>erives from rapi<em>d</em> association an<em>d</em> extremely slow <em>d</em>issociation of OASS with SAT an<em>d</em> requires the C terminus of SAT for the interaction. Stea<em>d</em>y-state kinetic analysis shows that CSC formation enhances SAT activity an<em>d</em> releases SAT from substrate inhibition an<em>d</em> fee<em>d</em>back inhibition by cysteine, the final pro<em>d</em>uct of the biosynthesis pathway. Cysteine inhibits SAT an<em>d</em> the CSC with K(i) values of <em>2</em> an<em>d</em> 70 microm, respectively. These results suggest a new mo<em>d</em>el for the architecture of this regulatory complex an<em>d</em> a<em>d</em><em>d</em>itional control mechanisms for biochemically controlling plant cysteine biosynthesis. Base<em>d</em> on previous work an<em>d</em> our results, we suggest that OASS acts as an enzyme chaperone of SAT in the CSC.

The equilibrium mixture of yeast enolase with substrate, <em>2</em>-phospho-<em>D</em>-glycerate (<em>2</em>-PGA), and product, phosphoenolpyruvate (P-enolpyruvate), has been crystallized from solutions of poly(ethylene glycol) (PEG) at pH 8.0. Crystals belong to the space group C<em>2</em> and have unit cell dimensions a = 1<em>2</em>1.9 A, b = 73.<em>2</em> A, c = 93.9 A, and beta = 93.3 degrees. The crystals have one <em>dimer</em> per asymmetric unit. Crystals of the equilibrium mixture and of the enolase complex of phosphonoacetohydroxamate (PhAH) are isomorphous, and the structure of the former complex was solved from the coordinates of enolase-(Mg<em>2</em>+)<em>2</em>-PhAH [Wedekind, J. E., Poyner, R. R., Reed, G. H., & Rayment, I. (1994) Biochemistry 33, 9333-934<em>2</em>]. The current crystallographic R-factor is 17.7% for all recorded data (9<em>2</em>% complete) to 1.8 A resolution. The electron density map is unambiguous with respect to the positions and liganding of both magnesium ions and with respect to the stereochemistry of substrate/product binding. Both magnesium ions are complexed to functional groups of the substrate/product. The higher affinity Mg<em>2</em>+ coordinates to the carboxylate side chains of Asp <em>2</em>46, Glu <em>2</em>95, and Asp 3<em>2</em>0, both carboxylate oxygens of the substrate/product, and a water molecule. One of the carboxylate oxygens of the substrate/product also coordinates to the lower affinity Mg<em>2</em>+-thus forming a mu-carboxylato bridge. The other ligands of the second Mg<em>2</em>+ are a phosphoryl oxygen of the substrate/product, two water molecules, and the carbonyl and gamma-oxygens of Ser 39 from the active site loop. The intricate coordination of both magnesium ions to the carboxylate group suggests that both metal ions participate in stabilizing negative charge in the carbanion (aci-carboxylate) intermediate. The epsilon-amino group of Lys 345 is positioned to serve as the base in the forward reaction whereas the carboxylate side chain of Glu <em>2</em>11 is positioned to interact with the 3-OH of <em>2</em>-PGA. The structure provides a candid view of the catalytic machinery of enolase.

We have characterized the minimal functioning unit of UhpT, the secondary carrier that mediates exchange of phosphate and glucose 6-phosphate in Escherichia coli. Membranes of a UhpT overproducing strain were solubilized with 1.<em>2</em>5% octyl beta-<em>D</em>-glucopyranoside, in the presence of 0.1% E. coli phospholipid and with <em>2</em>0% glycerol as the osmolyte stabilant. That soluble UhpT could bind its natural substrates was indicated by the protections afforded by sugar phosphates against thermal inactivation or chemical modification with pyridoxal 5'-phosphate. Moreover, the degree of protection correlated with the strength of interaction between UhpT and the test substrate (<em>2</em>-deoxyglucose 6-phosphate = glucose 6-phosphate greater than galactose 6-phosphate = glucose 1-phosphate much greater than glucose 6-sulfate). Other experiments demonstrated that soluble UhpT existed as a monomer. For example, during both high performance liquid chromatography and conventional gel permeation chromatography, the elution pattern of UhpT activity was measured directly by a rapid reconstitution technique. In both cases, and in the presence and absence of substrate, UhpT activity traveled as a single component of Mr 53,000, corresponding closely to the sequence prediction of 50,600. Finally, reconstitution was studied at protein to lipid ratios low enough to achieve between 0.075 and 1.5 UhpT monomers/proteoliposome. Specific activity was constant throughout this range, a finding consistent with the idea of a functional monomer. Mitochondria and chloroplasts provide the only other anion exchange carriers described at this level of biochemical resolution, and these organelle antiporters function as <em>dimers</em>. By contrast, work summarized here places their bacterial counterpart, UhpT, in the same class as the lactose carrier of E. coli and the glucose carrier of the human erythrocyte, both of which function as monomers. Consideration of this pattern in conjunction with the known hydropathy profiles of these proteins suggests a novel scheme for the classification of all secondary carriers, with implications for both the structure and origin of these transport proteins.

Although mesenchymal stromal cells (MSCs) can promote wound healing in different clinical settings, the underlying mechanism of MSC-mediated tissue repair has yet to be determined. Because a nonredundant role of pentraxin 3 (PTX3) in tissue repair and remodeling has been recently described, here we sought to determine whether MSC-derived PTX3 might play a role in wound healing. Using a murine model of skin repair, we found that Ptx3-deficient (Ptx3(-/-)) MSCs delayed wound closure and reduced granulation tissue formation compared with wt MSCs. At day <em>2</em>, confocal microscopy revealed a dramatic reduction in green fluorescent protein (GFP)-expressing Ptx3(-/-) MSCs recruited to the wound, where they appeared to be not only poorly organized in bundles but also scattered in the extracellular matrix. These findings were further confirmed by quantitative biochemical analysis of GFP content in wound extracts. Furthermore, Ptx3(-/-) MSC-treated skins displayed increased levels of fibrin and lower levels of <em>D</em>-<em>dimer</em>, suggesting delayed fibrin-rich matrix remodeling compared with control skins. Consistently, both pericellular fibrinolysis and migration through fibrin were found to be severely affected in Ptx3(-/-) MSCs. Overall, our findings identify an essential role of MSC-derived PTX3 in wound repair underscoring the beneficial potential of MSC-based therapy in the management of intractable wounds.