R Hal Scofield

Oklahoma City University, Oklahoma City, Oklahoma, United States

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Publications (275)1201.02 Total impact

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    ABSTRACT: Objectives Following up the systemic lupus erythematosus (SLE) genome-wide association studies (GWAS) identification of NMNAT2 at rs2022013, we fine-mapped its 150 kb flanking regions containing NMNAT2 and SMG7 in a 15 292 case–control multi-ancestry population and tested functions of identified variants. Methods We performed genotyping using custom array, imputation by IMPUTE 2.1.2 and allele specific functions using quantitative real-time PCR and luciferase reporter transfections. SLE peripheral blood mononuclear cells (PBMCs) were cultured with small interfering RNAs to measure antinuclear antibody (ANA) and cyto/chemokine levels in supernatants using ELISA. Results We confirmed association at NMNAT2 in European American (EA) and Amerindian/Hispanic ancestries, and identified independent signal at SMG7 tagged by rs2702178 in EA only (p=2.4×10−8, OR=1.23 (95% CI 1.14 to 1.32)). In complete linkage disequilibrium with rs2702178, rs2275675 in the promoter region robustly associated with SMG7 mRNA levels in multiple expression quantitative trait locus (eQTL) datasets. Its risk allele was dose-dependently associated with decreased SMG7 mRNA levels in PBMCs of 86 patients with SLE and 119 controls (p=1.1×10−3 and 6.8×10−8, respectively) and conferred reduced transcription activity in transfected HEK-293 (human embryonic kidney cell line) and Raji cells (p=0.0035 and 0.0037, respectively). As a critical component in the nonsense-mediated mRNA decay pathway, SMG7 could regulate autoantigens including ribonucleoprotein (RNP) and Smith (Sm). We showed SMG7 mRNA levels in PBMCs correlated inversely with ANA titres of patients with SLE (r=−0.31, p=0.01), and SMG7 knockdown increased levels of ANA IgG and chemokine (C-C motif) ligand 19 in SLE PBMCs (p=2.0×10−5 and 2.0×10−4, respectively). Conclusion We confirmed NMNAT2 and identified independent SMG7 association with SLE. The inverse relationship between levels of the risk allele-associated SMG7 mRNAs and ANA suggested the novel contribution of mRNA surveillance pathway to SLE pathogenesis.
    No preview · Article · Jan 2016 · Annals of the Rheumatic Diseases
  • B.T. Kurien · R.H. Scofield
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    ABSTRACT: The transfer of biomolecules to a membrane support, referred to as blotting, has been an integral part of life sciences research. Southern first described the transfer of DNA to nitrocellulose membrane in 1975 (Southern blotting), spawning methods to transfer RNA (northern blotting) and proteins (western blotting). The transfer of protein patterns from a gel to a microporous membrane has been utilized universally to detect and characterize proteins, especially those of low abundance. Protein blotting has evolved greatly since its inception in 1979, as detailed in this work, allowing protein transfer to be accomplished in a variety of ways.
    No preview · Chapter · Dec 2015
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    ABSTRACT: Objective: More than 80% of autoimmune disease is female dominant, but the mechanism for this female bias is poorly understood. We suspected an X chromosome dose effect and hypothesized that trisomy X (47,XXX, 1 in ∼1,000 live female births) would be increased in female predominant diseases (e.g. systemic lupus erythematosus [SLE], primary Sjögren's syndrome [SS], primary biliary cirrhosis [PBC] and rheumatoid arthritis [RA]) compared to diseases without female predominance (sarcoidosis) and controls. Methods: We identified 47,XXX subjects using aggregate data from single nucleotide polymorphism (SNP) arrays and confirmed, when possible, by fluorescent in situ hybridization (FISH) or quantitative polymerase chain reaction (q-PCR). Results: We found 47,XXX in seven of 2,826 SLE and three of 1,033 SS female patients, but only in two of the 7,074 female controls (p=0.003, OR=8.78, 95% CI: 1.67-86.79 and p=0.02, OR=10.29, 95% CI: 1.18-123.47; respectively). One 47,XXX subject was present for ∼404 SLE women and ∼344 SS women. 47,XXX was present in excess among SLE and SS subjects. Conclusion: The estimated prevalence of SLE and SS in women with 47,XXX was respectively ∼2.5 and ∼2.9 times higher than in 46,XX women and ∼25 and ∼41 times higher than in 46,XY men. No statistically significant increase of 47,XXX was observed in other female-biased diseases (PBC or RA), supporting the idea of multiple pathways to sex bias in autoimmunity. This article is protected by copyright. All rights reserved.
    Full-text · Article · Dec 2015 · Arthritis and Rheumatology
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    ABSTRACT: Objectives: Autoantibodies reactive with Ro52 (tripartite motif (TRIM) containing protein 21 [TRIM21]) are detected in 70% of primary Sjögren's syndrome (pSjS) patients. TRIM21 belongs to a 34 member C-IV family of TRIM proteins. Although autoantibodies against other TRIM proteins within the C-IV family have been detected in the sera of pSjS patients, their clinical relevance remains unclear. This study investigates the frequency of anti-TRIM38 in pSjS patients and evaluates its association with different clinical measures of the disease. Methods: Serum samples from pSjS patients (n=235) and controls (n=50) were analyzed for reactivity to in vitro transcribed and translated (35) S-Met-TRIM38 protein. The association of anti-TRIM38 with different laboratory and clinical measures of pSjS were evaluated. Reactivity of anti-TRIM38 with different structural domains of TRIM38 was analyzed. Affinity purified anti-TRIM38 antibodies were used to immunoprecipitate TRIM21. Results: TRIM38 reactive autoantibodies were detected in the sera of 24/235 pSjS patients and in 2/50 controls. Anti-TRIM38 positivity was significantly associated with the presence of anti-Ro60, anti-Ro52, anti-La, rheumatoid factor and hypergammaglobulinemia. Clinically, anti-TRIM38 was associated with significantly higher ocular surface staining scores, lower Schirmer's test scores and minor labial salivary gland biopsy focus scores of ≥3.0. Anti-TRIM38 antibodies mainly recognized the Cortactin-binding protein-2 (CortBP-2, amino acids 128-238) and the B30.2/SPRY (amino acids 268-465) domains on TRIM38. Affinity purified antibodies to TRIM38-CortBP2 and TRIM38-B30.2/SPRY domains reacted with TRIM21. Conclusion: Our data demonstrate that anti-TRIM38 specificity arising in a subset of pSjS patients is associated with higher severity of pSjS. This article is protected by copyright. All rights reserved.
    No preview · Article · Dec 2015 · Arthritis and Rheumatology
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    ABSTRACT: Background/Purpose: There are currently no FDA approved immunomodulating agents available for treatment of the extraglandular manifestations of Sjogren's (SS). Clinical practice guidelines were developed for the rheumatologic/systemic manifestations of SS in order to improve quality and consistency of care.
    Full-text · Conference Paper · Nov 2015
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    Biji T Kurien · R Hal Scofield

    Full-text · Dataset · Nov 2015
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    ABSTRACT: Leptin is abnormally elevated in the plasma of patients with systemic lupus erythematosus (SLE), where it is thought to promote and/or sustain pro-inflammatory responses. Whether this association could reflect an increased genetic susceptibility to develop SLE is not known, and studies of genetic associations with leptin-related polymorphisms in SLE patients have been so far inconclusive. Here we genotyped DNA samples from 15,706 SLE patients and healthy matched controls from four different ancestral groups, to correlate polymorphisms of genes of the leptin pathway to risk for SLE. It was found that although several SNPs showed weak associations, those associations did not remain significant after correction for multiple testing. These data do not support associations between defined leptin-related polymorphisms and increased susceptibility to develop SLE.
    No preview · Article · Sep 2015 · Clinical Immunology
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    ABSTRACT: Objectives: Commercial curcumin (CU), derived from food spice turmeric (TU), has been widely studied as a potential therapeutic for a variety of oncological and inflammatory conditions. Lack of solubility/bioavailability has hindered curcumin's therapeutic efficacy in human diseases. We have solubilised curcumin in water applying heat/pressure, obtaining up to 35-fold increase in solubility (ultrasoluble curcumin (UsC)). We hypothesised that UsC or ultrasoluble turmeric (UsT) will ameliorate systemic lupus erythematosus (SLE) and Sjögren's syndrome (SS)-like disease in MRL-lpr/lpr mice. Methods: Eighteen female MRL-lpr/lpr (6 weeks old) and 18 female MRL-MpJ mice (6 weeks old) were used. Female MRL-lpr/lpr mice develop lupus-like disease at the 10th week and die at an average age of 17 weeks. MRL-MpJ mice develop lupus-like disease around 47 weeks and typically die at 73 weeks. Six mice of each strain received autoclaved water only (lpr-water or MpJ-water group), UsC (lpr-CU or MpJ-CU group) or UsT (lpr-TU or MpJ-TU group) in the water bottle. Results: UsC or UsT ameliorates SLE in the MRL-lpr/lpr mice by significantly reducing lymphoproliferation, proteinuria, lesions (tail) and autoantibodies. lpr-CU group had a 20% survival advantage over lpr-water group. However, lpr-TU group lived an average of 16 days shorter than lpr-water group due to complications unrelated to lupus-like illness. CU/TU treatment inhibited lymphadenopathy significantly compared with lpr-water group (p=0.03 and p=0.02, respectively) by induction of apoptosis. Average lymph node weights were 2606±1147, 742±331 and 385±68 mg, respectively, for lpr-water, lpr-CU and lpr-TU mice. Transferase dUTP nick end labelling assay showed that lymphocytes in lymph nodes of lpr-CU and lpr-TU mice underwent apoptosis. Significantly reduced cellular infiltration of the salivary glands in the lpr-TU group compared with the lpr-water group, and a trend towards reduced kidney damage was observed in the lpr-CU and lpr-TU groups. Conclusions: These studies show that UsC/UsT could prove useful as a therapeutic intervention in SLE/SS.
    Full-text · Article · Sep 2015
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    ABSTRACT: In recent years, a new class of drugs has revolutionized the treatment of autoimmune, allergic, infectious, and many more diseases. This new class of drugs is made of 3 groups-cytokines, monoclonal antibodies, and fusion proteins-that may target special damaged cells but not all the cells. These drugs may have side effects such as infection, hypersensitivity, hematologic disorders, cancer, hepatotoxicity, and neurologic disorders. However, there is not enough evidence or long-term studies of the mechanism of action and side effects of these drugs. Patients receiving biological therapies may need special consideration in dentistry. This paper is a review of the classification, mechanism of action, and side effects of these drugs and dental consideration for patients receiving biological therapies.
    No preview · Article · Sep 2015
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    Biji T Kurien · R Hal Scofield
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    ABSTRACT: Several techniques have been employed to detect proteins on membranes. These include the use of quantum dot luminescent labels, oxyblot immunochemical detection, polymer immunocomplexes, "coupled" probing approach, in situ renaturation of proteins for detecting enzyme activities in crude or purified preparations, immunochromatographic assay, western-phosphatase assay, and use of Congo red dye (a cosmetic color named Alta), Pro-Q Emerald 488 dye, or amine-reactive dye in combination with alkaline phosphatase- and horseradish peroxidase-antibody conjugates for the simultaneous trichromatic fluorescence detection of proteins. Several methods have been used to improve the detection of proteins on membranes, including glutaraldehyde treatment of nitrocellulose blots, elimination of keratin artifacts in immunoblots probed with polyclonal antibodies, and washing of immunoblots with excessive water and manipulation of Tween-20 in wash buffer. These methods are briefly reviewed in this chapter.
    Full-text · Article · Jul 2015 · Methods in molecular biology (Clifton, N.J.)
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    Biji T Kurien · R Hal Scofield
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    ABSTRACT: Minor impurities in tryptic peptide digests can affect the signal obtained in matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Therefore, it becomes necessary to purify the digests, especially those that fail to yield good mass spectra. Here, we describe a simple protocol using polyvinylidene difluoride membrane for purifying tryptic peptides prior to mass spectrometric analysis. The tryptic digest is spotted on a polyvinylidene difluoride membrane, air-dried, and washed. The membrane is then extracted with trifluoroacetic acid/acetonitrile and the extract is then subjected to matrix-assisted laser desorption ionization time-of-flight mass spectrometry. This method enabled us to identify a cross-reactive D1 autoantigen on the surface of neutrophils that bound antibodies targeting Ro 60 autoantigen in systemic lupus erythematosus.
    Full-text · Article · Jul 2015 · Methods in molecular biology (Clifton, N.J.)
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    Biji T Kurien · Debashish Danda · Robert H Scofield

    Full-text · Article · Jul 2015 · International Journal of Rheumatic Diseases
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    ABSTRACT: To characterise the clinical features, immunological profile and outcome in a cohort of Asian Indian patients with primary Sjögren's syndrome (SS). Electronic medical records from a tertiary care teaching hospital in south India were screened for SS between 2004 and 2011. Patients fulfilling American European Consensus group (AECG) 2002 or American College of Rheumatology (ACR) 2012 classification criteria were included. Agglomerative hierarchical cluster analysis to identify patterns of associations between clinical and immunological features was done. Multivariate logistic regression to identify predictors of major systemic involvement was performed. Data on treatment and outcome were retrieved from electronic records. Of 423 patients suspected to have SS, 332 fulfilled inclusion criteria. Only 8.3% of patients complained of sicca symptoms on their own at initial presentation. Younger age of onset, higher female to male ratio, paucity of cryoglobulinemia, Raynaud's phenomenon and hyperglobulinemia were unique to this cohort. Cluster analysis revealed two subsets: The first cluster comprised of patients having a major systemic illness with high antibody titers and the second comprised of seronegative patients with mild disease. Over a third of SS cases had severe systemic manifestations necessitating treatment with immunosuppressants. In multivariate logistic regression analysis, anti-Ro and anti-La antibody positivity was associated with higher odds for systemic disease features (OR=2.67, P=0.03 and OR=3.25, P=0.003, respectively) whereas chronic pain was associated with lower odds (OR=0.4, p=0.032). Clinical improvement including symptomatic benefit in sicca and musculoskeletal features was noted with immunomodulators in the majority. Our cohort of patients with SS has characteristic clinical features; some of them are in contrast with previous observations reported in European patients. This cohort consisted of two distinct patient clusters. The first cluster was associated with major systemic illness and high antibody titers, where as the second cluster comprised of seronegative patients with mild disease. Association of antibody positivity with systemic features was further confirmed on logistic regression analysis.
    Full-text · Article · Jun 2015 · The Open Rheumatology Journal
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    Biji T Kurien · Debashish Danda · R Hal Scofield
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    ABSTRACT: Dactyloscopy or fingerprint identification is a vital part of forensic evidence. Identification with fingerprints has been known since the finding of finger impressions on the clay surface of Babylonian legal contracts almost 4,000 years ago. The skin on the fingers and palms appears as grooves and ridges when observed under a microscope. A unique fingerprint is produced by the patterns of these friction skin ridges. Visible fingerprints can be deposited on solid surfaces. Colored inks have been used to deposit fingermarks on documents. Herein, we show that alkaline phosphatase can be used to transfer prints from fingers or palm to nitrocellulose or polyvinylidene difluoride membranes. The prints can be detected by using the nitro blue tetrazolium/5-bromo-4-chloro-3-indolyl phosphate method of detection.
    Full-text · Article · Jun 2015 · Methods in molecular biology (Clifton, N.J.)
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    ABSTRACT: An ultra-rapid method for electrophoresing proteins on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transfer of proteins to nitrocellulose membranes, and immunoblotting is described here. Electrophoresis of the autoantigens La and Ro60, as well as molecular weight standards on a 4-20 % gradient gel, was performed in about 10 min using heated (70-75 °C) normal Laemmli running buffer. Electrophoretic transfer of these proteins was achieved in 7 min using a semidry transfer method. Finally, immunoblotting of La and Ro60 was carried out in 30 min. Thus, the entire process of electrophoresis, electrotransfer, and immunoblotting could be carried out in 1 h.
    Full-text · Article · Jun 2015 · Methods in molecular biology (Clifton, N.J.)
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    Biji T Kurien · R Hal Scofield
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    ABSTRACT: Western blotting enables the detection and characterization of proteins of low abundance. Sodium dodecyl sulfate (SDS) polyacrylamide gel-separated proteins are normally transferred electrophoretically to nitrocellulose or polyvinylidene difluoride membranes. Here we describe the transfer proteins [Ro 60 (or SSA) autoantigen, 220 and 240 kDa spectrin antigens, and prestained molecular weight standards] by diffusion from SDS polyacrylamide gels at 37 °C. Up to 12 immunoblots can be obtained from a single gel by this method.
    Full-text · Article · Jun 2015 · Methods in molecular biology (Clifton, N.J.)
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    Biji T Kurien · R Hal Scofield
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    ABSTRACT: A method for the electrophoretic transfer of high and low molecular weight proteins to nitrocellulose membranes following sodium dodecyl sulfate (SDS) polyacrylamide gel is described here. The transfer was performed with heated (70-75 °C) normal transfer buffer from which methanol had been omitted. Complete transfer of high and low molecular weight antigens (molecular weight protein standards, a purified protein, and proteins from a human tissue extract) could be carried out in 10 min for a 7 % (0.75 mm) SDS polyacrylamide gel. For 10 and 12.5 % gels (0.75 mm) the corresponding time was 15 min. A complete transfer could be carried out in 20 min for 7, 10, and 12.5 % gels (1.5 mm gels). The permeability of the gel is increased by heat, such that the proteins trapped in the polyacrylamide gel matrix can be easily transferred to the membrane. The heat mediated transfer method was compared with a conventional transfer protocol, under similar conditions. The conventional method transferred minimal low molecular weight proteins while retaining most of the high molecular weight proteins in the gel. In summary, this procedure is particularly useful for the transfer of high molecular weight proteins, very rapid, and avoids the use of methanol.
    Full-text · Article · Jun 2015 · Methods in molecular biology (Clifton, N.J.)
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    Biji T Kurien · R Hal Scofield
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    ABSTRACT: Proteins have been transferred from the gel to the membrane by a variety of methods. These include vacuum blotting, centrifuge blotting, electroblotting of proteins to Teflon tape and membranes for N- and C-terminal sequence analysis, multiple tissue blotting, a two-step transfer of low- and high-molecular-weight proteins, acid electroblotting onto activated glass, membrane-array method for the detection of human intestinal bacteria in fecal samples, protein microarray using a new black cellulose nitrate support, electrotransfer using square wave alternating voltage for enhanced protein recovery, polyethylene glycol-mediated significant enhancement of the immunoblotting transfer, parallel protein chemical processing before and during western blot and the molecular scanner concept, electronic western blot of matrix-assisted laser desorption/ionization mass spectrometric-identified polypeptides from parallel processed gel-separated proteins, semidry electroblotting of peptides and proteins from acid-urea polyacrylamide gels, transfer of silver-stained proteins from polyacrylamide gels to polyvinylidene difluoride (PVDF) membranes, and the display of K(+) channel proteins on a solid nitrocellulose support for assaying toxin binding. The quantification of proteins bound to PVDF membranes by elution of CBB, clarification of immunoblots on PVDF for transmission densitometry, gold coating of nonconductive membranes before matrix-assisted laser desorption/ionization tandem mass spectrometric analysis to prevent charging effect for analysis of peptides from PVDF membranes, and a simple method for coating native polysaccharides onto nitrocellulose are some of the methods involving either the manipulation of membranes with transferred proteins or just a passive transfer of antigens to membranes. All these methods are briefly reviewed in this chapter.
    Full-text · Article · Jun 2015 · Methods in molecular biology (Clifton, N.J.)
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    Biji T Kurien · R Hal Scofield
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    ABSTRACT: Western blotting is an important procedure for the immunodetection of proteins, particularly proteins that are of low abundance. This process involves the transfer of protein patterns from gel to microporous membrane. Electrophoretic as well as non-electrophoretic transfer of proteins to membranes was first described in 1979. Protein blotting has evolved greatly since the inception of this protocol, allowing protein transfer to be accomplished in a variety of ways.
    Full-text · Article · Jun 2015 · Methods in molecular biology (Clifton, N.J.)
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    Biji T Kurien · R Hal Scofield
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    ABSTRACT: A sensitive and specific method to analyze specific antibody clonotype changes in a lupus patient who developed autoantibodies to the Ro 60 autoantigen under observation is described. Patient sera, collected over several years, were separated by flatbed isoelectric focusing (IEF) and analyzed by affinity immunoblotting utilizing Ro 60-coated nitrocellulose membrane. When the Ro 60-coated nitrocellulose was laid over the surface of the IEF gel, the antibodies present on the surface of the acrylamide gel bound the Ro antigen on the nitrocellulose. Tween-20 was used to prevent nonspecific binding. The bound IgG clonotypes were detected using alkaline phosphatase conjugated anti-IgG. The patient's sera demonstrated an oligoclonal response to the Ro 60 autoantigen that increased in complexity and affinity over time.
    Full-text · Article · Jun 2015 · Methods in molecular biology (Clifton, N.J.)

Publication Stats

7k Citations
1,201.02 Total Impact Points

Institutions

  • 1999-2015
    • Oklahoma City University
      Oklahoma City, Oklahoma, United States
  • 1997-2015
    • University of Oklahoma
      Norman, Oklahoma, United States
  • 1991-2015
    • University of Oklahoma Health Sciences Center
      • • College of Medicine
      • • Department of Pathology
      • • Department of Biochemistry and Molecular Biology
      • • Section on Diabetes/Endocrinology
      Oklahoma City, Oklahoma, United States
  • 2014
    • University of Texas at El Paso
      El Paso, Texas, United States
  • 2003-2014
    • United States Department of Veterans Affairs
      Бедфорд, Massachusetts, United States
  • 1992-2014
    • Oklahoma Medical Research Foundation
      • Arthritis and Clinical Immunology Program
      Oklahoma City, Oklahoma, United States
  • 2012
    • University of Texas Southwestern Medical Center
      • Center for Cancer Immunobiology
      Dallas, Texas, United States
  • 2009
    • Carl Gustav Carus-Institut
      Pforzheim, Baden-Württemberg, Germany